Detection of clinically relevant copy number variants with whole-exome sequencing.
Copy number variation (CNV) is a common source of genetic variation that has been implicated in many genomic disorders. This has resulted in the widespread application of genomic microarrays as a first-tier diagnostic tool for CNV detection. More recently, whole-exome sequencing (WES) has been proven successful for the detection of clinically relevant point mutations and small insertion-deletions exome wide. We evaluate the utility of short-read WES (SOLiD 5500xl) to detect clinically relevant CNVs in DNA from 10 patients with intellectual disability and compare these results to data from two independent high-resolution microarrays. Eleven of the 12 clinically relevant CNVs were detected via read-depth analysis of WES data; a heterozygous single-exon deletion remained undetected by all algorithms evaluated. Although the detection power of WES for small CNVs currently does not match that of high-resolution microarray platforms, we show that the majority (88%) of rare coding CNVs containing three or more exons are successfully identified by WES. These results show that the CNV detection resolution of WES is comparable to that of medium-resolution genomic microarrays commonly used as clinical assays. The combined detection of point mutations, indels, and CNVs makes WES a very attractive first-tier diagnostic test for genetically heterogeneous disorders.


Streamlining Safety Data Collection in Hospital-Acquired Bacterial Pneumonia and Ventilator-Associated Bacterial Pneumonia Trials: Recommendations of the Clinical Trials Transformation Initiative Antibacterial Drug Development Project Team.
Resistant bacteria are one of the leading causes of hospital-acquired/ventilator-associated bacterial pneumonia (HABP/VABP). HABP/VABP trials are complex and difficult to conduct due to the large number of medical procedures, adverse events, and concomitant medications involved. Differences in the legislative frameworks between different regions of the world may also lead to excessive data collection. The Clinical Trials Transformation Initiative (CTTI) seeks to advance antibacterial drug development (ABDD) by streamlining clinical trials to improve efficiency and feasibility while maintaining ethical rigor, patient safety, information value, and scientific validity. In 2013, CTTI engaged a multidisciplinary group of experts to discuss challenges impeding the conduct of HABP/VABP trials. Separate workstreams identified challenges associated with current data collection processes. Experts defined "data collection" as the act of capturing and reporting certain data on the case report form as opposed to recording of data as part of routine clinical care. The ABDD Project Team developed strategies for streamlining safety data collection in HABP/VABP trials using a Quality by Design approach. Current safety data collection processes in HABP/VABP trials often include extraneous information. More targeted strategies for safety data collection in HABP/VABP trials will rely on optimal protocol design and prespecification of which safety data are essential to satisfy regulatory reporting requirements. A consensus and a cultural change in clinical trial design and conduct, which involve recognition of the need for more efficient data collection, are urgently needed to advance ABDD and to improve HABP/VABP trials in particular.


Merging dietary assessment with the adolescent lifestyle.
The use of image-based dietary assessment methods shows promise for improving dietary self-report among children. The Technology Assisted Dietary Assessment (TADA) food record application is a self-administered food record specifically designed to address the burden and human error associated with conventional methods of dietary assessment. Users would take images of foods and beverages at all eating occasions using a mobile telephone or mobile device with an integrated camera [e.g. Apple iPhone, Apple iPod Touch (Apple Inc., Cupertino, CA, USA); Nexus One (Google, Mountain View, CA, USA)]. Once the images are taken, the images are transferred to a back-end server for automated analysis. The first step in this process is image analysis (i.e. segmentation, feature extraction and classification), which allows for automated food identification. Portion size estimation is also automated via segmentation and geometric shape template modeling. The results of the automated food identification and volume estimation can be indexed with the Food and Nutrient Database for Dietary Studies to provide a detailed diet analysis for use in epidemiological or intervention studies. Data collected during controlled feeding studies in a camp-like setting have allowed for formative evaluation and validation of the TADA food record application. This review summarises the system design and the evidence-based development of image-based methods for dietary assessment among children.


A randomized trial comparing treatments for varicose veins.
Ultrasound-guided foam sclerotherapy and endovenous laser ablation are widely used alternatives to surgery for the treatment of varicose veins, but their comparative effectiveness and safety remain uncertain. In a randomized trial involving 798 participants with primary varicose veins at 11 centers in the United Kingdom, we compared the outcomes of foam, laser, and surgical treatments. Primary outcomes at 6 months were disease-specific quality of life and generic quality of life, as measured on several scales. Secondary outcomes included complications and measures of clinical success. After adjustment for baseline scores and other covariates, the mean disease-specific quality of life was slightly worse after treatment with foam than after surgery (P=0.006) but was similar in the laser and surgery groups. There were no significant differences between the surgery group and the foam or the laser group in measures of generic quality of life. The frequency of procedural complications was similar in the foam group (6%) and the surgery group (7%) but was lower in the laser group (1%) than in the surgery group (P<0.001); the frequency of serious adverse events (approximately 3%) was similar among the groups. Measures of clinical success were similar among the groups, but successful ablation of the main trunks of the saphenous vein was less common in the foam group than in the surgery group (P<0.001). Quality-of-life measures were generally similar among the study groups, with the exception of a slightly worse disease-specific quality of life in the foam group than in the surgery group. All treatments had similar clinical efficacy, but complications were less frequent after laser treatment and ablation rates were lower after foam treatment. (Funded by the Health Technology Assessment Programme of the National Institute for Health Research; Current Controlled Trials number, ISRCTN51995477.).


Lower hand grip strength in older adults with non-alcoholic fatty liver disease: a nationwide population-based study.
Although both liver and muscle are metabolically active endocrine organs, and non-alcoholic fatty liver disease (NAFLD) and sarcopenia may share common pathogenic determinants, there have been few clinical studies of the relationship between NAFLD and muscle strength, especially in the elderly. We conducted a nationally representative population-based, cross-sectional study using data from the Korea National Health and Nutrition Examination Survey, which involved 1,897 men aged ≥50 years and 2,206 postmenopausal women. NAFLD was defined using the hepatic steatosis index (HSI) and low muscle strength was defined using the Korea-specific cut-off point of hand grip strength (HGS). Men and women with NAFLD had 7.3% and 7.9% lower HGS than controls, respectively. The odds ratios for low muscle strength in the presence of NAFLD were 2.51 in men and 2.34 in women. HSI inversely correlated with HGS in both men and women. Consistently, compared with men and women in the lowest HSI quartile, those in the highest quartile had 7.6% and 12.4% lower HGS, respectively, and were 5.63- and 3.58-times more likely to have low muscle strength, respectively. These results provide the first clinical evidence that NAFLD can be associated with muscular impairment in older adults, as demonstrated by lower muscle strength.


Genetic diversity in butterflies: Interactive effects of habitat fragmentation and climate-driven range expansion.
Some species are expanding their ranges polewards during current climate warming. However, anthropogenic fragmentation of suitable habitat is affecting expansion rates and here we investigate interactions between range expansion, habitat fragmentation and genetic diversity. We examined three closely related Satyrinae butterflies, which differ in their habitat associations, from six sites along a transect in England from distribution core to expanding range margin. There was a significant decline in allozyme variation towards an expanding range margin in Pararge aegeria, which has the most restricted habitat availability, but not in Pyronia tithonus whose habitat is more widely available, or in a non-expanding 'control species' (Maniola jurtina). Moreover, data from another transect in Scotland indicated that declines in genetic diversity in P. aegeria were evident only on the transect in England, which had greater habitat fragmentation. Our results indicate that fragmentation of breeding habitats leads to more severe founder events during colonization, resulting in reduced diversity in marginal populations in more specialist species. The continued widespread loss of suitable habitats in the future may increase the likelihood of loss of genetic diversity in expanding species, which may affect whether or not species can adapt to future environmental change.


Sorting of the yeast vacuolar-type, proton-translocating ATPase enzyme complex (V-ATPase): identification of a necessary and sufficient Golgi/endosomal retention signal in Stv1p.
Subunit a of the yeast vacuolar-type, proton-translocating ATPase enzyme complex (V-ATPase) is responsible for both proton translocation and subcellular localization of this highly conserved molecular machine. Inclusion of the Vph1p isoform causes the V-ATPase complex to traffic to the vacuolar membrane, whereas incorporation of Stv1p causes continued cycling between the trans-Golgi and endosome. We previously demonstrated that this targeting information is contained within the cytosolic, N-terminal portion of V-ATPase subunit a (Stv1p). To identify residues responsible for sorting of the Golgi isoform of the V-ATPase, a random mutagenesis was performed on the N terminus of Stv1p. Subsequent characterization of mutant alleles led to the identification of a short peptide sequence, W(83)KY, that is necessary for proper Stv1p localization. Based on three-dimensional homology modeling to the Meiothermus ruber subunit I, we propose a structural model of the intact Stv1p-containing V-ATPase demonstrating the accessibility of the W(83)KY sequence to retrograde sorting machinery. Finally, we characterized the sorting signal within the context of a reconstructed Stv1p ancestor (Anc.Stv1). This evolutionary intermediate includes an endogenous W(83)KY sorting motif and is sufficient to compete with sorting of the native yeast Stv1p V-ATPase isoform. These data define a novel sorting signal that is both necessary and sufficient for trafficking of the V-ATPase within the Golgi/endosomal network.


Development and validation of the Geriatric Anxiety Inventory.
Anxiety symptoms and anxiety disorders are highly prevalent among elderly people, although infrequently the subject of systematic research in this age group. One important limitation is the lack of a widely accepted instrument to measure dimensional anxiety in both normal old people and old people with mental health problems seen in various settings. Accordingly, we developed and tested of a short scale to measure anxiety in older people. We generated a large number of potential items de novo and by reference to existing anxiety scales, and then reduced the number of items to 60 through consultation with a reference group consisting of psychologists, psychiatrists and normal elderly people. We then tested the psychometric properties of these 60 items in 452 normal old people and 46 patients attending a psychogeriatric service. We were able to reduce the number of items to 20. We chose a 1-week perspective and a dichotomous response scale. Cronbach's alpha for the 20-item Geriatric Anxiety Inventory (GAI) was 0.91 among normal elderly people and 0.93 in the psychogeriatric sample. Concurrent validity with a variety of other measures was demonstrated in both the normal sample and the psychogeriatric sample. Inter-rater and test-retest reliability were found to be excellent. Receiver operating characteristic analysis indicated a cut-point of 10/11 for the detection of DSM-IV Generalized Anxiety Disorder (GAD) in the psychogeriatric sample, with 83% of patients correctly classified with a specificity of 84% and a sensitivity of 75%. The GAI is a new 20-item self-report or nurse-administered scale that measures dimensional anxiety in elderly people. It has sound psychometric properties. Initial clinical testing indicates that it is able to discriminate between those with and without any anxiety disorder and between those with and without DSM-IV GAD.


Psychological and behavioral factors associated with colorectal cancer screening among Ashkenazim.
Psychological and behavioral factors related to annual colorectal cancer (CRC) screening were examined in a sample of Ashkenazi Jewish individuals. Identification of factors related to regular CRC screening in this population is important because of the possibility of a heightened incidence of CRC. Eligible participants were 171 Ashkenazi Jewish adults 40 years or older attending an educational program about breast cancer genetics. Compliance with recommended guidelines for digital rectal examination and fecal occult blood test in the past year were dependent measures. Demographic variables, family history of CRC, perceived risk, physician recommendation, and worry about cancer were independent measures. Digital rectal examinations and fecal occult blood tests had been obtained in the past year by 46 and 31% of the participants, respectively. A logistic regression showed that physician recommendation was related significantly to obtaining digital rectal examinations. Physician recommendation and education were related significantly to obtaining fecal occult blood tests. Although participants with family histories of CRC perceived themselves as being at increased risk of developing CRC, and were more worried about developing colon cancer, they were no more likely to adhere to CRC screening guidelines than those without such histories. Overall, compliance with recommended CRC screening was low even among high-risk individuals. Physicians play a key role in motivating people to comply with CRC screening. Physicians need to en courage all asymptomatic patients 50 years and older to be screened for CRC.


Acute lower respiratory infections on lung sequelae in Cambodia, a neglected disease in highly tuberculosis-endemic country.
Little is known about post-infectious pulmonary sequelae in countries like Cambodia where tuberculosis is hyper-endemic and childhood pulmonary infections are highly frequent. We describe the characteristics of hospitalized Cambodian patients presenting with community-acquired acute lower respiratory infections (ALRI) on post-infectious pulmonary sequelae (ALRIPS). Between 2007 and 2010, inpatients ≥15 years with ALRI were prospectively recruited. Clinical, biological, radiological and microbiological data were collected. Chest radiographs were re-interpreted by experts to compare patients with ALRIPS, on previously healthy lungs (ALRIHL) and active pulmonary tuberculosis (TB). Patients without chest radiograph abnormality or with abnormality suggestive as other chronic respiratory diseases were excluded from this analysis. Among the 2351 inpatients with community-acquired ALRI, 1800 were eligible: 426 (18%) ALRIPS, 878 (37%) ALRIHL and 496 (21%) TB. ALRIPS patients had less frequent fever than other ALRI (p < 0.001) and more productive cough than ALRIHL (p < 0.001). Streptococcus pneumoniae, Haemophilus influenzae, and Pseudomonas aeruginosa accounted for 83% of ALRIPS group positive cultures. H. influenzae and P. aeruginosa were significantly associated with ALRIPS compared with ALRIHL. Treatment was appropriate in 58% of ALRIPS patients. Finally, 79% of ALRIPS were not recognized by local clinicians. In-hospital mortality was low (1%) but probably underestimated in the ALRIPS group. ALRIPS remains often misdiagnosed as TB with inappropriate treatment in low-income countries. Better-targeted training programs would help reduce the morbidity burden and financial costs.


Molecular evolution in space and through time: mtDNA phylogeography of the Olive Sunbird (Nectarinia olivacea/obscura) throughout continental Africa.
This study constitutes the first investigation of the phylogeographic structure of a forest bird distributed throughout the montane and lowland forest biomes of Africa. The key objective was to investigate the importance of Pleistocene climatic cycles on avian diversification across Africa. The Olive Sunbird is a relatively large polytypic sunbird widely distributed throughout evergreen, montane and coastal forests in Africa. Recently, it was split into two species, the Eastern Olive Sunbird (Nectarinia olivacea) and the Western Olive Sunbird (Nectarinia obscura), based on morphological grounds. Analyses of a 395bp fragment of the mtDNA NADH subunit 3 gene with flanking tRNA sequences, from 196 individuals of N. olivacea and 86 from N. obscura indicate that genetic divergence levels are low (1.0-2.4%) across some 9000km, from Ghana in the northwest of Africa to KwaZulu-Natal in eastern South Africa. Neither currently recognized Olive Sunbird species were monophyletic using either parsimony or likelihood tree-building methods. Phi(ST) values suggested that there was less variation partitioned among species than between most neighboring regions. Genetic diversity within the N. olivacea/obscura complex was dominated by three star-like phylogenies linked to each other by a single mutational step and two subnetworks (IV and V) separated from the core star-like phylogenies (subnetworks I, II, and III) by five to six mutational steps. The dominant evolutionary mechanism shaping genetic variation within the N. olivacea/obscura complex as identified by nested-clade analyses, appears to be one of range expansion possibly out of East Africa associated with a period of forest expansion during the mid-Pleistocene, some 1.1-0.7 million years ago. Mismatch profiles suggested that secondary contact has occurred between eastern and western lineages within the Ufipa Escarpment and possibly Zimbabwe, as well as between eastern lineages in the Kenyan Highlands and northern Eastern Arc Mts.


Studies of the extracellular glycocalyx of the anaerobic cellulolytic bacterium Ruminococcus albus 7.
Anaerobic cellulolytic bacteria are thought to adhere to cellulose via several mechanisms, including production of a glycocalyx containing extracellular polymeric substances (EPS). As the compositions and structures of these glycocalyces have not been elucidated, variable-pressure scanning electron microscopy (VP-SEM) and chemical analysis were used to characterize the glycocalyx of the ruminal bacterium Ruminococcus albus strain 7. VP-SEM revealed that growth of this strain was accompanied by the formation of thin cellular extensions that allowed the bacterium to adhere to cellulose, followed by formation of a ramifying network that interconnected individual cells to one another and to the unraveling cellulose microfibrils. Extraction of 48-h-old whole-culture pellets (bacterial cells plus glycocalyx [G] plus residual cellulose [C]) with 0.1 N NaOH released carbohydrate and protein in a ratio of 1:5. Boiling of the cellulose fermentation residue in a neutral detergent solution removed almost all of the adherent cells and protein while retaining a residual network of adhering noncellular material. Trifluoroacetic acid hydrolysis of this residue (G plus C) released primarily glucose, along with substantial amounts of xylose and mannose, but only traces of galactose, the most abundant sugar in most characterized bacterial exopolysaccharides. Linkage analysis and characterization by nuclear magnetic resonance suggested that most of the glucosyl units were not present as partially degraded cellulose. Calculations suggested that the energy demand for synthesis of the nonprotein fraction of EPS by this organism represents only a small fraction (<4%) of the anabolic ATP expenditure of the bacterium.


Regulation of iron pathways in response to hypoxia.
Constituting an integral part of a heme's porphyrin ring, iron is essential for supplying cells and tissues with oxygen. Given tight links between oxygen delivery and iron availability, it is not surprising that iron deprivation and oxygen deprivation (hypoxia) have very similar consequences at the molecular level. Under hypoxia, the expression of major iron homeostasis genes including transferrin, transferrin receptor, ceruloplasmin, and heme oxygenase-1 is activated by hypoxia-inducible factors to provide increased iron availability for erythropoiesis in an attempt to enhance oxygen uptake and delivery to hypoxic cells. Iron-response proteins (IRP1 and IRP2) and "cap-n-collar" bZIP transcriptional factors (NE-F2 p45; Nrf1, 2, and 3; Bach1 and 2) also control gene and protein expression of the key iron homeostasis proteins. In this article, we give an overview of the mechanisms by which iron pathways are regulated by hypoxia at multiple levels. In addition, potential clinical benefits of manipulating iron pathways in the hypoxia-related conditions anemia and ischemia are discussed.


Polarity control of h-BN nanoribbon edges by strain and edge termination.
We studied the polarity of h-BN nano-flakes in terms of their edge geometries, edge hydrogen termination, and uniaxial strain by evaluating their electrostatic potential using density functional theory. Our calculations have shown that the polarity of the nanoribbons is sensitive to their edge shape, edge termination, and uniaxial tensile strain. Polarity inversion of the ribbons can be induced by controlling the hydrogen concentration at the edges and the uniaxial tensile strain. The polarity inversion indicates that h-BN nanoribbons can exhibit non-polar properties at a particular edge hydrogen concentration and tensile strain, even though the nanoribbons essentially have polarity at the edge. We also found that the edge angle affects the polarity of nanoribbons with hydrogenated edges.


Detection of intracellular granularity induction in prostate cancer cell lines by small molecules using the HyperCyt high-throughput flow cytometry system.
Prostate cancer is a leading cause of death among men due to the limited number of treatment strategies available for advanced disease. Discovery of effective chemotherapeutics involves the identification of agents that inhibit cancer cell growth. Increases in intracellular granularity have been observed during physiological processes that include senescence, apoptosis, and autophagy, making this phenotypic change a useful marker for identifying small molecules that induce cellular growth arrest or death. In this regard, epithelial-derived cancer cell lines appear uniquely susceptible to increased intracellular granularity following exposure to chemotherapeutics. We have established a novel flow cytometry approach that detects increases in side light scatter in response to morphological changes associated with intracellular granularity in the androgen-sensitive LNCaP and androgen-independent PC3 human prostate cancer cell lines. A cell-based assay was developed to screen for small molecule inducers of intracellular granularity using the HyperCyt high-throughput flow cytometry platform. Validation was performed using the Prestwick Chemical Library, where known modulators of LNCaP intracellular granularity, such as testosterone, were identified. Nonandrogenic inducers of granularity were also detected. A further screen of approximately 25,000 small molecules led to the identification of a class of aryl-oxazoles that increased intracellular granularity in both cell lines, often leading to cell death. The most potent agents exhibited submicromolar efficacy in LNCaP and PC3 cells.


The immunotoxic effects of dual exposure to PCP and TCDD.
Pentachlorophenol (PCP) was a commonly used fungicide, herbicide, insecticide, and bactericide in industrial, agricultural, and domestic settings; however, it was also contaminated with polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). It has been reported that technical grade PCP had immunosuppressive effects and that the immune system was the major target of PCDD/PCDFs toxicity. Although the immune response after exposure to PCP or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been studied, the toxic effects of exposure to both PCP and TCDD have not yet been reported. The aim of this study was to evaluate the effects on immune cells from mice intraperitoneally immunized with OVA and subsequently treated with PCP or TCDD alone or in combination by gavage. The animals were terminated on day 7 and 14, and the spleen and plasma samples were collected for immunotoxicity evaluation. The numbers and populations of splenocytes, T cell-derived cytokines produced by splenocytes, splenocyte-generated cytotoxicity and OVA-specific antibodies in plasma were investigated. Our results indicate that the spleen/body weight ratio and splenocyte number was reduced by TCDD alone; in addition, this reduction was enhanced when TCDD was combined with PCP. Exposure to TCDD alone or in conjunction with PCP suppressed many ovalbumin (OVA)-stimulated cytokines, including IL-2, IFN-γ, IL-4, IL-5, and IL-10. Furthermore, the immunoglobulins IgG and IgM were suppressed in mice administered by PCP alone, but the suppressive effects were greater in mice treated with TCDD alone or in combination with PCP. Co-exposure to PCP and TCDD resulted in an antagonistic effect on TCDD-induced suppression of IFN-γ and IL-10. Our results demonstrate that PCP alone is immunotoxic, regardless of the presence of TCDD. PCP led to mild changes in cytokine secretion, and it compromised splenocyte-generated cytotoxicity and IgM and IgG antibody production on day 7. The finding that PCP antagonizes TCDD-induced IFN-γ suppression could be due to the competitive binding of PCP to AhR (aryl hydrocarbon receptor).


[Effects on the lipid profile in humans of a polyphenol-rich carob (Ceratonia siliqua L.) extract in a dairy matrix like a functional food; a pilot study].
The design of functional foods enriched in nutrients that favorably alter the lipid profile to prevent cardiovascular diseases and stimulate bowel function is of great interest. We have assayed a non-extractable-tannates-rich carob-fiber (PF-1®) in a milk matrix developed by Biosearch S.A. to discover its effects on the lipid profile and bowel function of human volunteers. A 4-week interventional study (400 mL daily consumption of this functional food, containing 20 g of PF-1®/L), was conducted: blood samples were analyzed for lipid profile, glucose, transaminases, creatinine and fat-soluble vitamins. The body-mass index and bowel function of the participants in the study were also measured. A tendency for triglyceride levels to diminish was observed in all participants (P = 0.066), and in the normal-cholesterol group in particular (P = 0.078). Another tendency to total cholesterol levels fell in the hypercholesterolemic group (P = 0.061) was also found. In the normal-cholesterol group, total cholesterol (CT), HDL-cholesterol and LDL-cholesterol levels significantly increased with the consumption of the functional food (P < 0.05). A better bowel function was also recorded by volunteers. This preliminary study highlights the possible positive influence of this functional food on the regulation of the lipid profile and bowel function in humans.


The ability of phased-array MRI in preoperative staging of primary rectal cancer: correlation with histopathological results.
This study evaluated the accuracy of phased-array magnetic resonance imaging (MRI) for preoperative local tumor staging in primary rectal cancer and emphasized the importance of the preoperative differentiation of T2 tumors from T3 tumors so the appropriate treatment can be applied. Twenty-four patients with primary rectal cancer were examined preoperatively using 1.5 T MRI with a phased-array coil. Multiplanar T2-weighted images were obtained. Rectum anatomy, depth of tumor invasion, mesorectal involvement and lymph nodes were assessed. All patients underwent radical surgery. The histological sections were evaluated microscopically. The correlation of magnetic resonance imaging and histopathology was assessed using the kappa statistic. Overstaging with MRI was compared with Fischer's exact test. Histopathological examination of the tumors revealed adenocarcinoma. When the tumors were staged, there was one patient with a pT1 tumor, six patients with pT2 tumors, and 17 patients with pT3 tumors. Using MRI, four patients with pT2 were overstaged as T3, and one patient with pT3 was overstaged as T4. In the remaining cases (one pT1, two pT2, and 16 pT3), MRI correctly assessed the stage of transmural invasion. The accuracy of T staging and metastatic lymph node detection with MRI was calculated as 79.2% and 58.5%, respectively. Phased-array MRI is a valuable technique for the preoperative staging of rectal cancer, especially in the differentiation of T2 and T3 tumors.


The active site base controls cofactor reactivity in Escherichia coli amine oxidase: x-ray crystallographic studies with mutational variants.
Amine oxidases utilize a proton abstraction mechanism following binding of the amine substrate to the C5 position of the cofactor, the quinone form of trihydroxyphenylalanine (TPQ). Previous work [Wilmot, C. M., et al. (1997) Biochemistry 36, 1608-1620] has shown that Asp383 in Escherichia coliamine oxidase (ECAO) is the catalytic base which performs the key step of proton abstraction. This paper explores in more depth this and other roles of Asp383. The crystal structures of three mutational variants are presented together with their catalytic properties, visible spectra, and binding properties for a substrate-like inhibitor, 2-hydrazinopyridine (2-HP), in comparison to those of the wild type enzyme. In wild type ECAO, the TPQ is located in a wedge-shaped pocket which allows more freedom of movement at the substrate binding position (C5) than for TPQ ring carbons C1-C4. A role of Asp383, whose carboxylate is located close to O5, is to stabilize the TPQ in its major conformation in the pocket. Replacement of Asp383 with the isostructural, but chemically distinct, Asn383 does not affect the location or dynamics of the TPQ cofactor significantly, but eliminates catalytic activity and drastically reduces the affinity for 2-HP. Removal of the side chain carboxyl moiety, as in Ala383, additionally allows the TPQ the greater conformational flexibility to coordinate to the copper, which demonstrates that Asp383 helps maintain the active site structure by preventing TPQ from migrating to the copper. Glu383 has a greatly decreased catalytic activity, as well as a decreased affinity for 2-HP relative to that of wild type ECAO. The electron density reveals that the longer side chain of Glu prevents the pivotal motion of the TPQ by hindering its movement within the wedge-shaped active site pocket. The results show that Asp383 performs multiple roles in the catalytic mechanism of ECAO, not only in acting as the active site base at different stages of the catalytic cycle but also in regulating the mobility of the TPQ that is essential to catalysis.


Reproductive and sexual health benefits in private health insurance plans in Washington State.
Although unintended pregnancy and sexually transmitted diseases (STDs) are considerable problems in the United States, private health insurance plans are inconsistent in their coverage of reproductive and sexual health services needed to address these problems. A survey administered to a market-representative sample of 12 health insurance carriers in Washington State assessed benefit coverage for gynecologic services, maternity services, contraceptive services, pregnancy termination, infertility services, reproductive cancer screening, STD services, HIV and AIDS services, and sterilization, as well as for the existence of confidentiality policies. "Core" services in each category were defined based on U.S. Preventive Services Task Force and other recommendations. Of the 91 top-selling plans on which data were collected, 8% were indemnity plans, 14% were point-of-service plans, 21% were preferred-provider organization plans and 57% were health maintenance organization (HMO)-type products; they had a combined enrollment of 1.4 million individuals. Coverage of core services varied widely by type of plan. While a high proportion of plans covered core gynecologic, maternity, reproductive cancer screening, STD and HIV and AIDS services, nearly half of plans did not cover any kind of contraceptive method. Approximately 13% of female enrollees did not have core coverage for gyneco!ogic services, 19% for matemity services, 75% for contraception, 37% for sterilization and 53% for pregnancy termination; 98% of women and men were not covered for infertility treatment. Most carriers did not have specific policies for maintaining privacy of sensitive health information. Overall, benefit coverage was lower for indemnity, preferred-provider organization and HMO plans in Washington State than has previously been seen nationally. A sizable proportion of women and men in Washington State who rely on private-sector health insurance lack comprehensive coverage for key reproductive and sexual health services.


Detection, prevalence, and transmission of avian hematozoa in waterfowl at the Arctic/sub-Arctic interface: co-infections, viral interactions, and sources of variation.
The epidemiology of avian hematozoa at high latitudes is still not well understood, particularly in sub-Arctic and Arctic habitats, where information is limited regarding seasonality and range of transmission, co-infection dynamics with parasitic and viral agents, and possible fitness consequences of infection. Such information is important as climate warming may lead to northward expansion of hematozoa with unknown consequences to northern-breeding avian taxa, particularly populations that may be previously unexposed to blood parasites. We used molecular methods to screen blood samples and cloacal/oropharyngeal swabs collected from 1347 ducks of five species during May-August 2010, in interior Alaska, for the presence of hematozoa, Influenza A Virus (IAV), and IAV antibodies. Using models to account for imperfect detection of parasites, we estimated seasonal variation in prevalence of three parasite genera (Haemoproteus, Plasmodium, Leucocytozoon) and investigated how co-infection with parasites and viruses were related to the probability of infection. We detected parasites from each hematozoan genus in adult and juvenile ducks of all species sampled. Seasonal patterns in detection and prevalence varied by parasite genus and species, age, and sex of duck hosts. The probabilities of infection for Haemoproteus and Leucocytozoon parasites were strongly positively correlated, but hematozoa infection was not correlated with IAV infection or serostatus. The probability of Haemoproteus infection was negatively related to body condition in juvenile ducks; relationships between Leucocytozoon infection and body condition varied among host species. We present prevalence estimates for Haemoproteus, Leucocytozoon, and Plasmodium infections in waterfowl at the interface of the sub-Arctic and Arctic and provide evidence for local transmission of all three parasite genera. Variation in prevalence and molecular detection of hematozoa parasites in wild ducks is influenced by seasonal timing and a number of host traits. A positive correlation in co-infection of Leucocytozoon and Haemoproteus suggests that infection probability by parasites in one or both genera is enhanced by infection with the other, or that encounter rates of hosts and genus-specific vectors are correlated. Using size-adjusted mass as an index of host condition, we did not find evidence for strong deleterious consequences of hematozoa infection in wild ducks.


Mycobacterial Infections in Rabbits: From the Wild to the Laboratory.
Tuberculous mycobacterial diseases such as leprosy and tuberculosis are ancient diseases that currently continue threatening human health in some countries. Non-tuberculous mycobacterial (NTM) infections cause a series of well-defined pathological entities, as well as some opportunistic diseases that have also increased worldwide, being more common among immunocompromised patients but rising also in immunocompetent individuals. Reports on natural infections by mycobacteria in rabbits are scarce and mainly involve NTM such as Mycobacterium avium subsp. avium in pigmy rabbits in the United States and Mycobacterium avium subsp. paratuberculosis in wild rabbits in Europe. Rabbits have been used as laboratory animals through the years, both to generate immunological reagents and as infection models. Mycobacterial infection models have been developed in this animal species showing different susceptibility patterns to mycobacteria in laboratory conditions. The latent tuberculosis model and the cavitary tuberculosis model have been widely used to elucidate pathogenic mechanisms and to evaluate chemotherapy and vaccination strategies. Rabbits have also been used as bovine paratuberculosis infection models. This review aimed to gather both wildlife and experimental infection data on mycobacteriosis in rabbits to assess their role in the spread of these infections as well as their potential use in the experimental study of mycobacterial pathogenesis and treatment.


Activation of endothelial cell phospholipase D by sphingosine and sphingosine-1-phosphate.
We have investigated the activation of phospholipase D (PLD) by sphingosine and its derivatives in bovine pulmonary artery endothelial cells (BPAEC) prelabeled with [32P]orthophosphate or [32P]lyso phospholipids. Sphingosine, in a dose- and time-dependent manner, stimulated the hydrolysis of [32P]phosphatidylcholine (PC) resulting in the production of [32P]phosphatidic acid (PA), suggesting PLD activation. In the presence of ethanol (150 mM), the accumulation of [32P]phosphatidylethanol was also observed. The sphingosine-induced stimulation of PLD activity was not affected by treatment with the protein kinase C (PKC) inhibitor staurosporine or by down-regulation of PKC with TPA and was independent of extracellular Ca2+, suggesting that the PLD activation was independent of PKC and Ca2+. Chelation of intracellular Ca2+ with BAPTA actually potentiated the sphingosine-stimulated [32P]PC hydrolysis. Furthermore, the activation of PLD by sphingosine was not abolished by treatment of BPAEC with either cholera or pertussis toxin, indicating noninvolvement of toxin-sensitive G-proteins. In addition to hydrolysis of [32P]PC, sphingosine also stimulated PLD-mediated hydrolysis of [32P]phosphatidylethanolamine and [32P]phosphatidylinositol. Among the various sphingoid compounds, in addition to sphingosine, only sphingosine-1-phosphate (Sph-1-P) activated the endothelial cell PLD. The effect of sphingosine and Sph-1-P on PA phosphatase (PA Pase) activity was tested using [3H]glycerol-labeled PA. The Mg(2+)-independent and membrane-associated PA Pase activity was inhibited by sphingosine (IC50 = 200 microM) but not by Sph-1-P. This implies that sphingosine and Sph-1-P share a similar PLD-stimulating property but differ in their PA Pase inhibitory activity.


Extract of Magnoliae Flos inhibits ovariectomy-induced osteoporosis by blocking osteoclastogenesis and reducing osteoclast-mediated bone resorption.
Bone homeostasis is maintained by a balance between bone resorption by osteoclasts and bone formation by osteoblasts. Osteoporosis occurs when osteoclast activity surpasses osteoblast activity. Pro-inflammatory cytokines stimulate osteoclast differentiation and activity by increasing production of macrophage-colony stimulating factor and receptor activator of nuclear factor-κB ligand (RANKL). In this study, we investigated whether Magnoliae Flos (MF), one of the most commonly used Chinese medicinal herbs for managing rhinitis, sinusitis and headache, could effectively inhibit osteoporosis. In ovariectomized (OVX) mice compared to sham mice, the body weight increased and serum levels of alkaline phosphatase (ALP), tartrate resistant acid phosphatase 5b, calcium, and osteocalcin were significantly elevated. However, orally administrated MF extract substantially inhibited the increased body weight and serum levels of bone turnover markers, without any evidence of tissue toxicity. MF extract treatment significantly reversed the morphometric parameters of ovariectomy-induced bone loss, including trabecular bone volume, thickness, number, separation, and bone density, to almost the same levels of the sham mice. Furthermore, MF extract reduced the RANKL-mediated osteoclast differentiation and bone resorption by inhibiting the activities of matrix metalloproteinases (MMPs) and cathepsin K in mouse bone marrow macrophages. MF extract appeared to increase ALP activity in murine osteoblastic cells. Taken together, MF extract may be a beneficial supplement for the blockade of osteoporosis progression, particularly for the management of postmenopausal osteoporosis.


Use of a culture system to identify possible causes of abnormal retinal development.
The aim of this study was to apply a recently developed cell culture system to the problem of identifying possible etiologies of altered retinal maturation. Using techniques of electrophysiology and neuropharmacology, it was possible to monitor the release of acetylcholine at synapses formed by cultured retinal neurons derived from the fetal rat. The major finding of this study was that the functional development of cholinergic retinal neurons of the fetus could be altered if the mother rat had been injected with a synthetic glucocorticoid hormone or stressed by cold exposure during a critical period in pregnancy. Thus, the maturation of at least certain fetal retinal neurons appears to be influenced by factors affecting the mother. The culture system described here provides a potentially useful approach to the identification and investigation of possible causes of abnormalities in the functional development of the mammalian retina.


Size-assortative mating and sexual size dimorphism are predictable from simple mechanics of mate-grasping behavior.
A major challenge in evolutionary biology is to understand the typically complex interactions between diverse counter-balancing factors of Darwinian selection for size assortative mating and sexual size dimorphism. It appears that rarely a simple mechanism could provide a major explanation of these phenomena. Mechanics of behaviors can predict animal morphology, such like adaptations to locomotion in animals from various of taxa, but its potential to predict size-assortative mating and its evolutionary consequences has been less explored. Mate-grasping by males, using specialized adaptive morphologies of their forelegs, midlegs or even antennae wrapped around female body at specific locations, is a general mating strategy of many animals, but the contribution of the mechanics of this wide-spread behavior to the evolution of mating behavior and sexual size dimorphism has been largely ignored. Here, we explore the consequences of a simple, and previously ignored, fact that in a grasping posture the position of the male's grasping appendages relative to the female's body is often a function of body size difference between the sexes. Using an approach taken from robot mechanics we model coercive grasping of females by water strider Gerris gracilicornis males during mating initiation struggles. We determine that the male optimal size (relative to the female size), which gives the males the highest grasping force, properly predicts the experimentally measured highest mating success. Through field sampling and simulation modeling of a natural population we determine that the simple mechanical model, which ignores most of the other hypothetical counter-balancing selection pressures on body size, is sufficient to account for size-assortative mating pattern as well as species-specific sexual dimorphism in body size of G. gracilicornis. The results indicate how a simple and previously overlooked physical mechanism common in many taxa is sufficient to account for, or importantly contribute to, size-assortative mating and its consequences for the evolution of sexual size dimorphism.


Vitamin D and musculoskeletal health, cardiovascular disease, autoimmunity and cancer: Recommendations for clinical practice.
There is increasing evidence that, in addition to the well-known effects on musculoskeletal health, vitamin D status may be related to a number of non-skeletal diseases. An international expert panel formulated recommendations on vitamin D for clinical practice, taking into consideration the best evidence available based on published literature today. In addition, where data were limited to smaller clinical trials or epidemiologic studies, the panel made expert-opinion based recommendations. Twenty-five experts from various disciplines (classical clinical applications, cardiology, autoimmunity, and cancer) established draft recommendations during a 2-day meeting. Thereafter, representatives of all disciplines refined the recommendations and related texts, subsequently reviewed by all panelists. For all recommendations, panelists expressed the extent of agreement using a 5-point scale. Recommendations were restricted to clinical practice and concern adult patients with or at risk for fractures, falls, cardiovascular or autoimmune diseases, and cancer. The panel reached substantial agreement about the need for vitamin D supplementation in specific groups of patients in these clinical areas and the need for assessing their 25-hydroxyvitamin D (25(OH)D) serum levels for optimal clinical care. A target range of at least 30 to 40 ng/mL was recommended. As response to treatment varies by environmental factors and starting levels of 25(OH)D, testing may be warranted after at least 3 months of supplementation. An assay measuring both 25(OH)D(2) and 25(OH)D(3) is recommended. Dark-skinned or veiled individuals not exposed much to the sun, elderly and institutionalized individuals may be supplemented (800 IU/day) without baseline testing.


Modulation of platinum sensitivity and resistance by cyclosporin A in refractory ovarian and fallopian tube cancer patients: a phase II study.
Our objective was to assess the activity of cyclosporin A (CsA) used as a chemomodulator of carboplatin in refractory ovarian and fallopian tube cancer patients. Fifty-one patients (47 epithelial ovarian, 1 ovarian mixed mesodermal tumor, and 3 fallopian tube carcinomas) were enrolled in a prospective Phase II trial of CsA and carboplatin. CsA was infused as a loading dose of 10 mg/kg over 5 h, followed by carboplatin infused over 30 min at an AUC of 6 mg/ml x min, then a 24-h continuous infusion of 11.6 mg/kg CsA. The patients received this protocol as second- to sixth-line therapy and had received between 1 and 3 prior platinum-based regimens. Eight patients received more than six cycles every 28 days, 34 patients received three to six cycles; and 9 patients received only one or two cycles. Thirty-eight patients were evaluable for objective response, and in an additional nine patients, CA-125 was the only marker of response. Four patients had no marker of disease. Of evaluable patients, 74% were platinum resistant. There were nine objective responses (one complete and eight partial responses) for an overall response rate in evaluable patients of 24%, with a median duration of response of 7 months (range, 3-38+ months). No responses were seen in patients who had received only one or two cycles of therapy. Among the strictly defined platinum-resistant patients, there was an overall 14% response rate, including one partial response seen after five prior regimens of chemotherapy including paclitaxel, and one ongoing complete response for 38+ months. Among the rest of the patients (those who were potentially platinum sensitive), there was an overall 50+ response rate; four of five responses were seen in patients with a platinum-free interval of <24 months, with only one response seen in a patient with a platinum-free interval of >24 months. Of evaluable patients, 34% had stable disease for a duration of 3-19 months. The most common grade 3 or 4 toxicity, thrombocytopenia, was seen in 22% of the patients. Hypertension, which responded to medications, was seen in 18% of the patients during the CsA infusion. We concluded that this CsA/carboplatin regimen is active in potentially platinum-sensitive patients and compares well with the expected response rate of 30% in patients with a platinum-free interval <24 months who are retreated with platinum. Moreover, this regimen had modest but real activity in platinum-resistant patients.


Co-occurring mental health and alcohol misuse: dual disorder symptoms in combat injured veterans.
Service members face difficulties during military deployment potentially resulting in morbidities such as posttraumatic stress disorder (PTSD), depression, and alcohol misuse. The co-occurrence of alcohol misuse and mental health disorders is termed dual disorder and has been associated with adverse outcomes. The study included 812 high-risk (i.e., endorsing combat exposure with documented combat injury) male U.S. veterans of Operation Iraqi Freedom, injured between October 2004 and November 2007, identified from the Expeditionary Medical Encounter Database. PTSD and depression symptoms were significant correlates of alcohol misuse. Veterans with dual disorder symptoms reported a significantly higher mean number of health complaints on the Post-Deployment Health Reassessment compared with those endorsing only mental health symptoms. These results highlight how mental health disorders among injured service members increases the odds of problem drinking and those with dual disorder have elevated health complaints.


Copy number variants in schizophrenia: confirmation of five previous findings and new evidence for 3q29 microdeletions and VIPR2 duplications.
To evaluate previously reported associations of copy number variants (CNVs) with schizophrenia and to identify additional associations, the authors analyzed CNVs in the Molecular Genetics of Schizophrenia study (MGS) and additional available data. After quality control, MGS data for 3,945 subjects with schizophrenia or schizoaffective disorder and 3,611 screened comparison subjects were available for analysis of rare CNVs (<1% frequency). CNV detection thresholds were chosen that maximized concordance in 151 duplicate assays. Pointwise and genewise analyses were carried out, as well as analyses of previously reported regions. Selected regions were visually inspected and confirmed with quantitative polymerase chain reaction. In analyses of MGS data combined with other available data sets, odds ratios of 7.5 or greater were observed for previously reported deletions in chromosomes 1q21.1, 15q13.3, and 22q11.21, duplications in 16p11.2, and exon-disrupting deletions in NRXN1. The most consistently supported candidate associations across data sets included a 1.6-Mb deletion in chromosome 3q29 (21 genes, TFRC to BDH1) that was previously described in a mild-moderate mental retardation syndrome, exonic duplications in the gene for vasoactive intestinal peptide receptor 2 (VIPR2), and exonic duplications in C16orf72. The case subjects had a modestly higher genome-wide number of gene-containing deletions (>100 kb and >1 Mb) but not duplications. The data strongly confirm the association of schizophrenia with 1q21.1, 15q13.3, and 22q11.21 deletions, 16p11.2 duplications, and exonic NRXN1 deletions. These CNVs, as well as 3q29 deletions, are also associated with mental retardation, autism spectrum disorders, and epilepsy. Additional candidate genes and regions, including VIPR2, were identified. Study of the mechanisms underlying these associations should shed light on the pathophysiology of schizophrenia.


Detection of in vivo production of tumour necrosis factor-alpha by human thyroid epithelial cells.
We have established previously that human thyroid epithelial cells (TEC) from patients with autoimmune thyroiditis are able to synthesize cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6). This paper examines TEC in sections from autoimmune thyroiditis for the in vivo production of tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) using the combined techniques of in situ hybridization and immunohistochemistry. Thyroid tissue from patients with Graves' disease, Hashimoto's disease and non-toxic goitre was examined and both mRNA and the protein of TNF-alpha were detected in TEC on frozen sections. Representative figures of only Graves' samples are illustrated in this paper. In contrast, using the same methods, IFN-gamma was detected only in the infiltrating cells and not in TEC of thyroid tissue from the patients.


Help-Seeking Experiences of Immigrant Domestic Violence Survivors in Australia: A Snapshot of Muslim Survivors.
There has been significant research on women's experiences of domestic violence (DV) as well as on the women's help-seeking behaviors when living with partner abuse. Most of the Australian literature has focused on nonimmigrant women. We know that help seeking can include informal sources such as family, friends, religious leaders, and colleagues or formal assistance from police, doctors, social workers, counselors, and DV agencies. The current study aims to contribute to the literature on help seeking by looking at what has been found concerning immigrant DV survivors and complementing that with interview material from a sample of 14 Muslim immigrant DV survivors in New South Wales. First, we look at barriers that these women may encounter in seeking help and the non-help-seeking strategies they may employ. We then see what may trigger seeking help (including before and after leaving the abusers). Next, we look at how the two types of help seeking are used to better understand the positives and negatives of these pathways. This article ends with some suggestions for developing more appropriate and targeted strategies to assist abused immigrant DV survivors and their children.


IL-15 promotes activation and expansion of CD8+ T cells in HIV-1 infection.
In HIV-1-infected patients, increased numbers of circulating CD8+ T cells are linked to increased risk of morbidity and mortality. Here, we identified a bystander mechanism that promotes CD8 T cell activation and expansion in untreated HIV-1-infected patients. Compared with healthy controls, untreated HIV-1-infected patients have an increased population of proliferating, granzyme B+, CD8+ T cells in circulation. Vβ expression and deep sequencing of CDR3 revealed that in untreated HIV-1 infection, cycling memory CD8 T cells possess a broad T cell repertoire that reflects the repertoire of the resting population. This suggests that cycling is driven by bystander activation, rather than specific antigen exposure. Treatment of peripheral blood mononuclear cells with IL-15 induced a cycling, granzyme B+ phenotype in CD8+ T cells. Moreover, elevated IL-15 expression in the lymph nodes of untreated HIV-1-infected patients correlated with circulating CD8+ T cell counts and was normalized in these patients following antiretroviral therapy. Together, these results suggest that IL-15 drives bystander activation of CD8+ T cells, which predicts disease progression in untreated HIV-1-infected patients and suggests that elevated IL-15 may also drive CD8+ T cell expansion that is linked to increased morbidity and mortality in treated patients.


Placebo Analgesia Reduces Costly Prosocial Helping to Lower Another Person's Pain.
Painkiller administration lowers pain empathy, but whether this also reduces prosocial behavior is unknown. In this preregistered study, we investigated whether inducing analgesia through a placebo painkiller reduced effortful helping. When given the opportunity to reduce the pain of another person, individuals experiencing placebo analgesia (n = 45 adults from Austria; 21 male, 24 female) made fewer prosocial choices at the lowest helping level and exerted less physical effort when helping, compared with controls whose pain sensitivity was unaltered (n = 45; 21 male, 24 female). Self-reported empathic unpleasantness positively correlated with prosocial choices across the whole sample. While not replicating group differences in empathy, a mediation analysis revealed that the level of unpleasantness to other people's pain fully mediated the effect of placebo analgesia on prosocial choices. Given the importance of prosociality for social cohesion, these findings have broad potential implications both for individuals under the influence of painkillers and for society at large.


Population genetic characteristics of the STR Loci D21S11 and FGA in eight diverse human populations.
A highly polymorphic multiplex short tandem repeat (STR) system composed of D21S11, FGA, and the sex-typing system amelogenin (AMG) has been used to investigate allele frequency distributions in two Canadian Caucasian samples (British Columbia and Alberta), three Canadian aboriginal populations (Coastal Salishans from British Columbia, Ojibwa from northern Ontario, and Cree from Saskatchewan), and three ethnic groups from Singapore (Chinese, Malays, and Asian Indians). Using the automated fluorescence detection approach on an ABD 373A DNA Sequencer, we distinguished 20 D21S11 and 22 FGA alleles with a nearly equal representation of two- and four-base variants. An overlap in allele sizes for both STR loci across populations was observed, but frequency differences were noted. Statistical analysis revealed that (1) both D21S11 and FGA loci conform to Hardy-Weinberg equilibrium in all eight surveyed populations based on five different tests and (2) both STR loci are in linkage equilibrium. Results from the 2 x N contingency table exact tests for population differentiation demonstrated that the Canadian samples from two different provinces were not distinguishable from one another at either STR locus and therefore could be combined to form one Caucasian group. Likewise, Chinese and Malays from Singapore did not show significant differences at either STR locus. In contrast, all other examined populations exhibited differences deemed statistically significant. As a complement to our study, we compared D21S11 allele frequency distributions in 21 worldwide populations and FGA allele frequency distributions in 14 populations. Many alleles never previously reported in worldwide populations were identified in Canadian aboriginal and Asian samples from this study. Twenty-four D21S11 and 29 FGA alleles were distinguished in worldwide groups. Interesting similarities in allele frequency distribution patterns across populations suggest that the STR polymorphism at these loci predates the geographic dispersal of ancestral human populations. This study further demonstrates the utility of highly informative STR loci such as D21S11 and FGA in human population evolutionary history and in forensic medicine.


High plasma adiponectin levels in patients with severe traumatic brain injury.
Adiponectin plays an important role in the regulation of tissue inflammation. There is a paucity of data on circulating plasma adiponectin concentrations in human traumatic brain injury. This study is designed to investigate the potential associations between plasma adiponectin levels and clinical outcomes after traumatic brain injury. Plasma adiponectin levels of 86 patients with severe traumatic brain injury and 86 healthy subjects were determined. Clinical outcomes included in-hospital mortality, 6-month mortality and 6-month unfavorable outcome (Glasgow Outcome Scale score of 1-3). Plasma adiponectin levels were significantly higher in patients compared to controls (20.5±5.9 vs. 7.7±2.0μg/ml; P<0.001) and emerged as an independent predictor of in-hospital mortality [odds ratio (OR), 1.318; 95% confidence interval (CI), 1.049-1.629; P=0.008], 6-month mortality (OR, 1.328; 95% CI, 1.082-1.657; P=0.007) and 6-month unfavorable outcome (OR, 1.240; 95% CI, 1.066-1.443; P=0.005) in a multivariate analysis. For predicting these clinical outcomes, areas under receiver operating characteristic curve of plasma adiponectin level were similar to those of Glasgow Coma scale scores (all P>0.05). However, adiponectin did not improve predictive values of Glasgow Coma scale scores (all P>0.05). Plasma adiponectin level may represent a novel biomarker for predicting clinical outcomes of traumatic brain injury.


A prospective randomized controlled trial of the effects of vitamin D supplementation on cardiovascular disease risk.
Vitamin D (VitD) supplementation has been advocated for cardiovascular risk reduction; however, supporting data are sparse. The objective of this study was to determine whether VitD supplementation reduces cardiovascular risk. Subjects in this prospective, randomized, double-blind, placebo-controlled trial of post-menopausal women with serum 25-hydroxyvitamin D concentrations >10 and <60 ng/mL were randomized to Vitamin D3 2500 IU or placebo, daily for 4 months. Primary endpoints were changes in brachial artery flow-mediated vasodilation (FMD), carotid-femoral pulse wave velocity (PWV), and aortic augmentation index (AIx). The 114 subjects were mean (standard deviation) 63.9 (3.0) years old with a 25-hydroxyvitamin D level of 31.3 (10.6) ng/mL. Low VitD (<30 ng/mL) was present in 47% and was associated with higher body-mass index, systolic blood pressure, glucose, CRP, and lower FMD (all p<0.05). After 4 months, 25-hydroxyvitamin D levels increased by 15.7 (9.3) ng/mL on vitamin D3 vs. -0.2 (6.1) ng/mL on placebo (p<0.001). There were no significant differences between groups in changes in FMD (0.3 [3.4] vs. 0.3 [2.6] %, p = 0.77), PWV (0.00 [1.06] vs. 0.05 [0.92] m/s, p = 0.65), AIx (2.7 [6.3] vs. 0.9 [5.6] %, p = 0.10), or CRP (0.3 [1.9] vs. 0.3 [4.2] mg/L, p = 0.97). Multivariable models showed no significant interactions between treatment group and low VitD status (<30 ng/mL) for changes in FMD (p = 0.65), PWV (p = 0.93), AIx (p = 0.97), or CRP (p = 0.26). In conclusion, VitD supplementation did not improve endothelial function, arterial stiffness, or inflammation. These observations do not support use of VitD supplementation to reduce cardiovascular disease risk.


A grounded theory study of homeopathic practitioners' perceptions and experiences of the homeopathic consultation.
Research into the homeopathic consultation has largely focused on patients' experiences, although the practitioner is a crucial component of the therapeutic context and may have an important part in optimizing health outcomes. Therefore the aim of this qualitative research was to gain an in-depth understanding of homeopathic practitioners' perceptions and experiences of the consultation. Medical and non-medical homeopaths were sampled from the registers of the Faculty and Society of Homeopaths. Two phases of data collection were employed. Phase 1 used in depth face-to-face interviews enabling the development of an initial model of the homeopathic consultation. Phase 2 involved observations of homeopathic consultations and practitioner reflective diaries in order to confirm, refute, or enlarge the model. Using the constant comparative method of grounded theory five main categories emerged, exploring the journey, finding the level, responding therapeutically, understanding self, and connecting, forming a model entitled "a theoretical model of a UK classical homeopathic consultation" which describes how homeopaths view and enact the consultation process. This study suggests that the process of identifying and prescribing the remedy is embedded in the consultation, highlighting the interconnectedness of the whole homeopathic consultation and aspects of the consultation that are unique and specific to homeopathy.


Carbohydrate Scaffolds for the Study of the Autism-associated Bacterium, Clostridium bolteae.
A large number of children in the autism spectrum disorder suffer from gastrointestinal (GI) conditions, such as constipation and diarrhea. Clostridium bolteae is a part of a set of pathogens being regularly detected in the stool samples of hosts affected by GI and autism symptoms. Accompanying studies have pointed out the possibility that such microbes affect behaviour through the production of neurotoxic metabolites in a so-called, gut-brain connection. As an extension of our Clostridium difficile polysaccharide (PS)-based vaccine research, we engaged in the discovery of C. bolteae surface carbohydrates. So far, studies revealed that C. bolteae produces a specific immunogenic PS capsule comprised of disaccharide repeating blocks of mannose (Manp) and rhamnose (Rhap) units: α-D-Manp-(1→[-4)-β-D-Rhap- (1→3)-α-D-Manp-(1→]n. For vaccinology and further immunogenic experiments, a method to produce C. bolteae PS conjugates has been developed, along with the chemical syntheses of the PS non-reducing end linkage, with D-Rha or L-Rha, α-D-Manp-(1→4)-α-D-Rhap- (1→O(CH2)5NH2 and α-D-Manp-(1→4)-α-L-Rhap-(1→O(CH2)5NH2, equipped with an aminopentyl linker at the reducing end for conjugation purposes. The discovery of C. bolteae PS immunogen opens the door to the creation of non-evasive diagnostic tools to evaluate the frequency and role of this microbe in autistic subjects and to a vaccine to reduce colonization levels in the GI tract, thus impeding the concentration of neurotoxins.


The genetic architecture of climatic adaptation of tropical cattle.
Adaptation of global food systems to climate change is essential to feed the world. Tropical cattle production, a mainstay of profitability for farmers in the developing world, is dominated by heat, lack of water, poor quality feedstuffs, parasites, and tropical diseases. In these systems European cattle suffer significant stock loss, and the cross breeding of taurine x indicine cattle is unpredictable due to the dilution of adaptation to heat and tropical diseases. We explored the genetic architecture of ten traits of tropical cattle production using genome wide association studies of 4,662 animals varying from 0% to 100% indicine. We show that nine of the ten have genetic architectures that include genes of major effect, and in one case, a single location that accounted for more than 71% of the genetic variation. One genetic region in particular had effects on parasite resistance, yearling weight, body condition score, coat colour and penile sheath score. This region, extending 20 Mb on BTA5, appeared to be under genetic selection possibly through maintenance of haplotypes by breeders. We found that the amount of genetic variation and the genetic correlations between traits did not depend upon the degree of indicine content in the animals. Climate change is expected to expand some conditions of the tropics to more temperate environments, which may impact negatively on global livestock health and production. Our results point to several important genes that have large effects on adaptation that could be introduced into more temperate cattle without detrimental effects on productivity.


Periodic patterns of actin turnover in lamellipodia and lamellae of migrating epithelial cells analyzed by quantitative Fluorescent Speckle Microscopy.
We measured actin turnover in lamellipodia and lamellae of migrating cells, using quantitative Fluorescent Speckle Microscopy. Lamellae disassembled at low rates from the front to the back. However, the dominant feature in their turnover was a spatially random pattern of periodic polymerization and depolymerization moving with the retrograde flow. Power spectra contained frequencies between 0.5 and 1 cycle/min. The spectra remained unchanged when applying Latrunculin A and Jasplakinolide in low doses, except that additional frequencies occurred beyond 1 cycle/min. Whereas Latrunculin did not change the rate of mean disassembly, Jasplakinolide halted it completely, indicating that the steady state and the dynamics of actin turnover are differentially affected by pharmacological agents. Lamellipodia assembled in recurring bursts at the leading edge and disassembled approximately 2.5 microm behind. Events of polymerization correlated spatially and temporally with transient formation of Arp2/3 clusters. In lamellae, Arp2/3 accumulation and polymerization correlated only spatially, suggesting an Arp2/3-independent mechanism for filament nucleation. To acquire these data we had to enhance the resolution of quantitative Fluorescent Speckle Microscopy to the submicron level. Several algorithmic advances in speckle image processing are described enabling the analysis of kinetic and kinematic activities of polymer networks at the level of single speckles.


Promissory accounts of personalisation in the commercialisation of genomic knowledge.
As part of personalised medicine emerging from the human genomics revolution, many websites now offer direct-to-consumer genetic testing. Here, we examine three personal genomics companies--Navigenics, deCODEme and 23andMe--each of which represents contrasting registers of 'personalisation'. We identify three distinctive registers in these websites: a paternalistic (medical) register; a translational (scientific) register and a democratic (consumerist) register. We explore in detail the rhetorical and discourse devices employed in these websites to assess how personalised healthcare is promised to the public. Promising information that will empower prevention of common complex diseases and ensure better quality of life is conflated with promising greater access to personal information. The presence and absence of scientific legitimacy is related to concerns about accuracy and validity on the one side, and fears of paternalism and elitism on the other. Nevertheless, a common strategy uniting these different styles of personalisation is consumer empowerment. Finally, we consider the tension between the drive of translational medicine to make human genomic research practically relevant, and the intrinsic uncertainties of scientific research and show how, in the commercial domain, future risks are transformed into discourses of promise by concealing these uncertainties.


Biosynthesis of plant volatiles: nature's diversity and ingenuity.
Plant volatiles (PVs) are lipophilic molecules with high vapor pressure that serve various ecological roles. The synthesis of PVs involves the removal of hydrophilic moieties and oxidation/hydroxylation, reduction, methylation, and acylation reactions. Some PV biosynthetic enzymes produce multiple products from a single substrate or act on multiple substrates. Genes for PV biosynthesis evolve by duplication of genes that direct other aspects of plant metabolism; these duplicated genes then diverge from each other over time. Changes in the preferred substrate or resultant product of PV enzymes may occur through minimal changes of critical residues. Convergent evolution is often responsible for the ability of distally related species to synthesize the same volatile.


The Montefiore community children's project: a controlled study of cognitive and emotional problems of homeless mothers and children.
This study compares the prevalence of emotional, academic, and cognitive impairment in children and mothers living in the community with those living in shelters for the homeless. In New York City, 82 homeless mothers and their 102 children, aged 6 to 11, recruited from family shelters were compared to 115 nonhomeless mothers with 176 children recruited from classmates of the homeless children. Assessments included standardized tests and interviews. Mothers in shelters for the homeless showed higher rates of depression and anxiety than did nonhomeless mothers. Boys in homeless shelters showed higher rates of serious emotional and behavioral problems. Both boys and girls in homeless shelters showed more academic problems than did nonhomeless children. Study findings suggest a need among homeless children for special attention to academic problems that are not attributable to intellectual deficits in either children or their mothers. Although high rates of emotional and behavioral problems characterized poor children living in both settings, boys in shelters for the homeless may be particularly in need of professional attention.


Perceived discrimination predicts elevated biological markers of inflammation among sexual minority adults.
Sexual minority (SM) adults (those who are lesbian, gay, or bisexual) consistently report more health problems compared to heterosexuals, and they tend to experience excess social stress. Although numerous studies have established links between social stress and clinical outcomes in SM adults, few studies have examined biological factors that may help explain how social stress leads to health disparities among SM adults. We used data from the Midlife in the United States Study (MIDUS) to examine whether two inflammatory markers that have been commonly associated with social stress-C-reactive protein (CRP) and interleukin-6 (IL-6)-differed by sexual orientation and whether any differences were explained by perceptions of discrimination. Participants self-identified as heterosexual (n = 1956) or lesbian, gay, or bisexual (n = 81). After controlling for age, gender, race, and education, SM individuals had higher CRP and IL-6 than heterosexuals on average and these differences were partially explained by perceptions of discrimination. Implications for inflammatory pathways as mechanisms related to SM health disparities and discrimination are discussed.


Physical and functional interaction of CARMA1 and CARMA3 with Ikappa kinase gamma-NFkappaB essential modulator.
CARMA proteins are scaffold molecules that contain a caspase recruitment domain and a membrane-associated guanylate kinase-like domain. CARMA1 plays a critical role in mediating activation of the NFkappaB transcription factor following antigen receptor stimulation of both B and T lymphocytes. However, the biochemical mechanism by which CARMA1 regulates activation of NFkappaB remains to be determined. Here we have shown that CARMA1 and CARMA3 physically associate with Ikappa kinase gamma/NFkappaB essential modulator (IkappaKgamma-NEMO) in lymphoid and non-lymphoid cells. CARMA1 participates to an inducible large molecular complex that contains IkappaKgamma/NEMO, Bcl10, and IkappaKalpha/beta kinases. Expression of the NEMO-binding region of CARMA3 exerts a dominant negative effect on Bcl10-mediated activation of NFkappaB. Thus, our results provide direct evidence for physical and functional interaction between CARMA and the IkappaK complex and offer a biochemical framework to understand the molecular activities controlled by CARMA-1, -2, and -3 and Bcl10.


Tracking Health Data Is Not Enough: A Qualitative Exploration of the Role of Healthcare Partnerships and mHealth Technology to Promote Physical Activity and to Sustain Behavior Change.
Despite the recent explosion of the mobile health (mHealth) industry and consumer acquisition of mHealth tools such as wearable sensors and applications (apps), limited information is known about how this technology can sustain health behavior change and be integrated into health care. The objective of the study was to understand potential users' views of mHealth technology, the role this technology may have in promoting individual activity goals aimed at improving health, and the value of integrating mHealth technology with traditional health care. Four focus groups were conducted with adults interested in sharing their views on how mHealth technology could support wellness programs and improve health. Participants (n=30) were enrolled from an employee population at an academic health institution. Qualitative thematic analysis was used to code transcripts and identify overarching themes. Our findings suggest that tracking health data alone may result in heightened awareness of daily activity, yet may not be sufficient to sustain use of mHealth technology and apps, which often have low reuse rates. Participants suggested that context, meaning, and health care partnerships need to be incorporated to engage and retain users. In addition to these findings, drivers for mHealth technology previously identified in the literature, including integration and control of health data were confirmed in this study. This study explores ways that mHealth technologies may be used to not only track data, but to encourage sustained engagement to achieve individual health goals. Implications of these findings include recommendations for mHealth technology design and health care partnership models to sustain motivation and engagement, allowing individuals to achieve meaningful behavior change.


Emotional and behavioural needs of children and adolescents with intellectual disabilities in an urban conurbation.
Over the past decade, increased attention has been paid to identifying and responding to the emotional and behavioural needs of children and adolescents with intellectual disability (ID). The aims of the present study were to add to this body of knowledge by identifying factors associated with emotional and behavioural needs among a sample of children with ID drawn from a large urban conurbation. Information was collected by postal questionnaire (or interview for family carers who did not have English as their first language) from teachers and from family carers of 615 children administratively identified as having ID (47% of all children with ID). Results indicated that: (1) the administrative prevalence of moderate but not severe ID was associated with social deprivation whereas the prevalence of severe but not moderate ID appeared to be associated with ethnicity; (2) 54% of children scored above the threshold on the Developmental Behaviour Checklist (DBC)-primary family carer, and 37% of children scored above the threshold on the DBC-teacher; (3) social deprivation, male gender, less severe ID and having fewer physical or sensory impairments were associated with antisocial and disruptive behaviour; and (4) more severe ID and additional impairments were associated with anxiety, communication disturbance, social relating and self-absorbed behaviours. These results identify a range of risk factors associated with behavioural and emotional problems experienced by children with ID.


Structural Changes of Human Serum Albumin (HSA) in Simulated Microgravity.
Nowadays, the biological effects of microgravity have been the subject of various experimental researches. Microgravity has been confirmed to affect biological systems. Furthermore, as a result of improvement in space technology for instance a manned mission to the moon, probabilities for human exposed to microgravity have incremented undoubtedly. The purpose of this study was to investigate the probable biological effects of microgravity on the human serum albumin (HSA) structure after 3 and 24 h exposure. It is worth mentioning that this is the first effort to investigate the structural alternations of HSA under simulated microgravity condition in biophysico-chemical terms thru different spectroscopic instruments. 2D clinostat was utilized for simulating microgravity. The UV-Vis, intrinsic and extrinsic fluorescence, dynamic light scattering (DLS) and circular dichroism (CD) spectra of 3.76 µM HSA in Tris-HCl buffer (pH 7.4, 0.1 M) and 3.76 µM HSA in Tris-HCl buffer (pH 7.4, 0.1 M) kept at simulated microgravity for 3 and 24h were verified. The UV-Visible, near-UV-CD and intrinsic fluorescence spectroscopy represented that microgravity can remarkably change the tertiary structure of HSA. Additionally, the ANS affinity for HSA incremented when the protein was exposed to simulate microgravity compared to unexposed HSA, which may possibly have appeared attributable to expansion of the structure of simulated HSA. Fluorescence quenching by acrylamide demonstrated higher stern-volmer constant for exposed HSA. The results of zeta potential and dynamic light scattering (DLS) experiments depicted that simulated microgravity cause raise in the surface charge and size of HSA. Far-UV CD data demonstrated that simulated microgravity did not perturb the secondary structures of the protein. Collectively, our results suggest that HSA after 24 h exposure to microgravity can exhibit a molten globule (MG) structure. This is the first report to demonstrate the molten globule state formation in microgravity condition. Results from this study could give knowledge to understand the role of gravity on protein folding process. In addition, this finding could help to find out safe limits for astronauts and space travelers and to develop adequate countermeasures against any harmful effects of microgravity.


The Transcription Factor ArcA Modulates Salmonella's Metabolism in Response to Neutrophil Hypochlorous Acid-Mediated Stress.
Salmonella Typhimurium, a bacterial pathogen with high metabolic plasticity, can adapt to different environmental conditions; these traits enhance its virulence by enabling bacterial survival. Neutrophils play important roles in the innate immune response, including the production of microbicidal reactive oxygen species (ROS). In addition, the myeloperoxidase in neutrophils catalyzes the formation of hypochlorous acid (HOCl), a highly toxic molecule that reacts with essential biomolecules, causing oxidative damage including lipid peroxidation and protein carbonylation. The bacterial response regulator ArcA regulates adaptive responses to oxygen levels and influences the survival of Salmonella inside phagocytic cells. Here, we demonstrate by whole transcriptomic analyses that ArcA regulates genes related to various metabolic pathways, enabling bacterial survival during HOCl-stress in vitro. Also, inside neutrophils, ArcA controls the transcription of several metabolic pathways by downregulating the expression of genes related to fatty acid degradation, lysine degradation, and arginine, proline, pyruvate, and propanoate metabolism. ArcA also upregulates genes encoding components of the oxidative pathway. These results underscore the importance of ArcA in ATP generation inside the neutrophil phagosome and its participation in bacterial metabolic adaptations during HOCl stress.


Fitness's moderation of the facilitative effect of acute exercise on cognitive flexibility in older women.
A recent study indicated that acute aerobic exercise improves cognitive flexibility in adults. The current study assessed age, habitual physical activity, and physical fitness as moderators of this improvement and examined whether the gains still exist an hour after the exercise session. The alternative-uses test, assessing cognitive flexibility, was administered individually to 20 older (age 63.67 +/- 3.55 yr) and 19 young (age 23.9 +/- 1.22) women before, immediately after, and an hour after a single moderate aerobic-exercise session. Results indicated significant improvement in cognitive flexibility in the older group immediately after the exercise but a decrease at the 1-hr follow-up. Further analysis indicated that physical fitness accounted for this improvement (R = -.622, p < .01). No such differences were observed in the young group. Further studies are needed to examine the duration of this effect, as well as the role of physical fitness as a moderator of it.


Healthcare databases in Europe for studying medicine use and safety during pregnancy.
The aim of this study was to describe a number of electronic healthcare databases in Europe in terms of the population covered, the source of the data captured and the availability of data on key variables required for evaluating medicine use and medicine safety during pregnancy. A sample of electronic healthcare databases that captured pregnancies and prescription data was selected on the basis of contacts within the EUROCAT network. For each participating database, a database inventory was completed. Eight databases were included, and the total population covered was 25 million. All databases recorded live births, seven captured stillbirths and five had full data available on spontaneous pregnancy losses and induced terminations. In six databases, data were usually available to determine the date of the woman's last menstrual period, whereas in the remainder, algorithms were needed to establish a best estimate for at least some pregnancies. In seven databases, it was possible to use data recorded in the databases to identify pregnancies where the offspring had a congenital anomaly. Information on confounding variables was more commonly available in databases capturing data recorded by primary-care practitioners. All databases captured maternal co-prescribing and a measure of socioeconomic status. This study suggests that within Europe, electronic healthcare databases may be valuable sources of data for evaluating medicine use and safety during pregnancy. The suitability of a particular database, however, will depend on the research question, the type of medicine to be evaluated, the prevalence of its use and any adverse outcomes of interest. © 2014 The Authors. Pharmacoepidemiology and Drug Safety published by John Wiley & Sons, Ltd.


How often do duplicated genes evolve new functions?
A recently duplicated gene can either fix a null allele (becoming a pseudogene) or fix an (advantageous) allele giving a slightly different function, starting it on the road to evolving a new function. Here we examine the relative probabilities of these two events under a simple model. Null alleles are assumed to be neutral; linkage effects are ignored, as are unequal crossing over and gene conversion. These assumptions likely make our results underestimates for the probability that an advantageous allele is fixed first. When new advantageous mutations are additive with selection coefficient s and the ratio of advantageous to null mutations is rho, the probability an advantageous allele is fixed first is ([1 - e-s]/[rho S] + 1)-1, where S = 4Nes with Ne the effective population size. The probability that a duplicate locus becomes a pseudogene, as opposed to evolving a new gene function, is high unless rhoS >> 1. However, even if advantageous mutations are very rare relative to null mutations, for sufficiently large populations rhoS >> 1 and new gene function, rather than pseudogene formation, is the expected fate of most duplicated genes.


The removal of carbohydrates from ricin with endoglycosidases H, F and D and alpha-mannosidase.
Recently, several investigators have explored the possibility of targeting ricin to designated cell types in animals by its linkage to specific antibodies. There is evidence, however, that the mannose-containing oligosaccharide chains on ricin are recognised by reticuloendothelial cells in the liver and spleen and so cause the immunotoxins to be removed rapidly from the blood stream. In the present study we analysed the carbohydrate composition of ricin and examined enzymic methods for removing the carbohydrate. The carbohydrate analysis ricin A-chain revealed the presence of one residue of xylose and one of fucose in addition to mannose and N-acetylglucosamine which had been detected previously. The B-chain contained only mannose and N-acetylglycosamine. Ricin A-chain is heterogeneous containing two components of molecular weight 30 000 and 32 000. Strong evidence was found that the heavier form of the A-chain contains an extra carbohydrate unit which is heterogeneous with respect to concanavalin A binding and sensitivity to endoglycosidase H. The lower molecular weight form of A-chain did not bind concanavalin A and was insusceptible to endoglycosidases. Only one of the two high mannose oligosaccharide units on the isolated B-chain could be removed by endoglycosidases H or F, whereas both were removable after denaturation of the polypeptide by SDS. Both the isolated A- and B-chains were sensitive to alpha-mannosidase. Intact ricin was resistant to endoglycosidase treatment and was only slightly sensitive to alpha-mannosidase. The addition of SDS allowed endoglycosidase H to remove both of the B-chain oligosaccharides from intact ricin and increased the toxin's sensitivity to alpha-mannosidase. In conclusion, extensive enzymic deglycosylation of ricin may only be possible if the A- and B-chains are first separated, treated with enzymes and then recombined to form the toxin.


Orientia tsutsugamushi: analysis of the mobilome of a highly fragmented and repetitive genome reveals ongoing lateral gene transfer in an obligate intracellular bacterium.
The rickettsial human pathogen Orientia tsutsugamushi (Ot) is an obligate intracellular Gram-negative bacterium with one of the most highly fragmented and repetitive genomes of any organism. Around 50% of its ~2.3 Mb genome is comprised of repetitive DNA that is derived from the highly proliferated Rickettsiales amplified genetic element (RAGE). RAGE is an integrative and conjugative element (ICE) that is present in a single Ot genome in up to 92 copies, most of which are partially or heavily degraded. In this report, we analysed RAGEs in eight fully sequenced Ot genomes and manually curated and reannotated all RAGE-associated genes, including those encoding DNA mobilisation proteins, P-type (vir) and F-type (tra) type IV secretion system (T4SS) components, Ankyrin repeat- and tetratricopeptide repeat-containing effectors, and other piggybacking cargo. Originally, the heavily degraded Ot RAGEs led to speculation that they are remnants of historical ICEs that are no longer active. Our analysis, however, identified two Ot genomes harbouring one or more intact RAGEs with complete F-T4SS genes essential for mediating ICE DNA transfer. As similar ICEs have been identified in unrelated rickettsial species, we assert that RAGEs play an ongoing role in lateral gene transfer within the Rickettsiales. Remarkably, we also identified in several Ot genomes remnants of prophages with no similarity to other rickettsial prophages. Together these findings indicate that, despite their obligate intracellular lifestyle and host range restricted to mites, rodents and humans, Ot genomes are highly dynamic and shaped through ongoing invasions by mobile genetic elements and viruses.


Schistosomiasis Drug Discovery in the Era of Automation and Artificial Intelligence.
Schistosomiasis is a parasitic disease caused by trematode worms of the genus Schistosoma and affects over 200 million people worldwide. The control and treatment of this neglected tropical disease is based on a single drug, praziquantel, which raises concerns about the development of drug resistance. This, and the lack of efficacy of praziquantel against juvenile worms, highlights the urgency for new antischistosomal therapies. In this review we focus on innovative approaches to the identification of antischistosomal drug candidates, including the use of automated assays, fragment-based screening, computer-aided and artificial intelligence-based computational methods. We highlight the current developments that may contribute to optimizing research outputs and lead to more effective drugs for this highly prevalent disease, in a more cost-effective drug discovery endeavor.


Theoretical evaluation of structural models of the S2 state in the oxygen evolving complex of Photosystem II: protonation states and magnetic interactions.
Protonation states of water ligands and oxo bridges are intimately involved in tuning the electronic structures and oxidation potentials of the oxygen evolving complex (OEC) in Photosystem II, steering the mechanistic pathway, which involves at least five redox state intermediates S(n) (n = 0-4) resulting in the oxidation of water to molecular oxygen. Although protons are practically invisible in protein crystallography, their effects on the electronic structure and magnetic properties of metal active sites can be probed using spectroscopy. With the twin purpose of aiding the interpretation of the complex electron paramagnetic resonance (EPR) spectroscopic data of the OEC and of improving the view of the cluster at the atomic level, a complete set of protonation configurations for the S(2) state of the OEC were investigated, and their distinctive effects on magnetic properties of the cluster were evaluated. The most recent X-ray structure of Photosystem II at 1.9 Å resolution was used and refined to obtain the optimum structure for the Mn(4)O(5)Ca core within the protein pocket. Employing this model, a set of 26 structures was constructed that tested various protonation scenarios of the water ligands and oxo bridges. Our results suggest that one of the two water molecules that are proposed to coordinate the outer Mn ion (Mn(A)) of the cluster is deprotonated in the S(2) state, as this leads to optimal experimental agreement, reproducing the correct ground state spin multiplicity (S = 1/2), spin expectation values, and EXAFS-derived metal-metal distances. Deprotonation of Ca(2+)-bound water molecules is strongly disfavored in the S(2) state, but dissociation of one of the two water ligands appears to be facile. The computed isotropic hyperfine couplings presented here allow distinctions between models to be made and call into question the assumption that the largest coupling is always attributable to Mn(III). The present results impose limits for the total charge and the proton configuration of the OEC in the S(2) state, with implications for the cascade of events in the Kok cycle and for the water splitting mechanism.


Development of a pharmacist-psychiatrist collaborative medication therapy management clinic.
To describe a needs assessment, practice description, practice innovation and reimbursement of a psychiatric pharmacist medication therapy management (MTM) clinic with related challenges and opportunities. An MTM clinic established in collaboration with the Outpatient Psychiatric Services (OPS) at the University of California San Diego (UCSD), under contract with the San Diego County Health and Human Services Agency Adult and Older Adult Mental Health Services (A/OAMHS). Two board-certified psychiatric pharmacists provided direct patient care using a collaborative practice protocol 3 days per week. Clinical services included pharmacotherapy management, laboratory monitoring, medication counseling, and psychosocial referrals to other providers. Payment to UCSD OPS for clinical services was based on a contract between the San Diego County A/OAMHS and the clinic. Two pharmacists co-managed mental health patients and billed for medication management based on face-to-face contact time (medication minutes) and documentation time with each patient. Number of patients comanaged, dropout rates, visit duration, and billed minutes. From May 2009 to December 2010, two pharmacists comanaged 68 patients, mean (± SD) age 48.6 ± 11.6 years, diagnosed with major depressive, schizophrenic, schizoaffective, and/or bipolar disorder. A total of 56 (82.3%) patients were clinically stable and remained on the pharmacist caseload, but 12 (17.6%) patients were lost to follow-up (10 lost contact, 1 moved, 1 expired). On average, patients had 7.7 patient visits , for 491 total visits (with an average of 26 minutes per visit) that were billed at a rate of $4.82 per minute for medication minutes, translating to $84,542.80. With provider education and appropriate physician champions, pharmacists are able to work collaboratively with psychiatrists in a mental health clinic.


Improved experimental method for the determination of optical constants by excitation of surface plasmon polaritons.
We propose an improved method for attenuated-total-reflection experiments in the Otto configuration. This new technique overcomes the difficulties that arise from the nonuniformity of the air coupling gap. Thus attenuated-total-reflection measurements, under ideal adapted conditions, can be made for a wide range of wavelengths. The determination of optical constants by a three-parameter least-square fit of the experimental data to Fresnel's theory always yields ambiguous results. We found that the ambiguity is caused by the loss of the phase angle in the reflectometry. Measurements for many air gap thicknesses in normal or extreme attenuated-total-reflection techniques, overcome this difficulty.


The Pf332 gene codes for a megadalton protein of Plasmodium falciparum asexual blood stages.
We characterized the Plasmodium falciparum antigen 332 (Ag332) which is specifically expressed during the asexual intraerythrocytic cycle of the parasite. The corresponding Pf332 gene has been located in the subtelomeric region of chromosome 11. Furthermore, it is present in all strains so far analyzed and shows marked restriction length fragment polymorphism. Partial sequence and restriction endonuclease digestion of cloned fragments revealed that the Pf332 gene is composed of highly degenerated repeats rich in glutamic acid. Mung been nuclease digestion and Northern blot analysis suggested that the Pf332 gene codes for a protein of about 700 kDa. These data were further confirmed by Western blot and immunoprecipitation of parasites extracts with an antiserum raised against a recombinant clone expressing part of the Ag332. Confocal immunofluorescence showed that Ag332 is translocated from the parasite to the surface of infected red blood cells within vesicle-like structures. In addition, Ag332 was detected on the surface of monkey erythrocytes infected with Plasmodium falciparum.