315 research outputs found

    Complex regulation of BpeEF-OprC mediated drug efflux in Burkholderia pseudomallei

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    2016 Spring.Includes bibliographical references.Burkholderia pseudomallei (Bp) is a Gram-negative bacillus and the etiologic agent of melioidosis, a multifaceted syndrome causing high mortality in tropical regions of the world. The bacteria is classified as a Tier-1 Select Agent due to the seriousness of infection, low infectious dose, lack of effective vaccine, and difficulty of treatment. Bp’s many acquired and intrinsic antimicrobial resistance determinants make the study of these factors vital to improving the efficacy of bi-phasic treatment currently used to treat melioidosis. This study examines one factor in particular: the BpeEF-OprC efflux pump, a member of the resistance-nodulation and cell division family of efflux proteins, and capable of extruding both trimethoprim and sulfamethoxazole. A combination of these compounds (co-trimoxazole) is the first line of eradication phase therapy, making BpeEF-OprC the most clinically important efflux pump encoded by Bp. In spite of this, little is understood of the regulation of bpeEF-oprC, other than it is controlled in part by two LysR family proteins, BpeS, and BpeT. We hypothesized that these regulatory proteins 1) exert their action(s) by interacting with bpeEF-oprC at a specific site within the bpeT-llpE-bpeEF-oprC intergenic region, 2) are capable of influencing transcription of additional operons, and 3) that mutations to these proteins altered ability to form multimers, thereby influencing their function as observed by increased co-trimoxazole resistance and bpeF transcript levels. In Aim I of the study, we identified the cis regulatory regions by which these proteins interact within the bpeT-llpE-bpeE intergenic region using a combination of 5’ deletion assays, S1 nuclease protection, fluorescent-linked oligo extension and electrophoretic mobility assays. With this information we were able to locate bpeT transcriptional start sites and promoter regions as well as binding sites for both BpeT and BpeS. In Aim II, we examined the function of BpeT and BpeS as trans regulatory factors of BpeEF-OprC through mutation and deletion of both genes in part I, and as global regulatory factors in part II. Through overexpression and qRT-PCR or MIC analysis of wild type and mutant forms of both genes, we observed that while BpeT is a direct transcriptional activator of bpeEF-oprC, BpeS is not. Additionally, mutation position in BpeS seems to play a role in the expression phenotype of bpeEF-oprC. However, these mutations do not influence the ability of BpeS or BpeT to form multimers, as we observed no change between wild type and mutant protein oligomer formation through low-pressure gel chromatography and native gel electrophoresis. These same mutations also appeared to have no deleterious effect on the ability of the protein to bind their consensus region within the IR. Additionally, the loss of both genes did not interrupt the ability of bpeEF-oprC to be induced by substrates of BpeEF-OprC, suggesting an additional regulatory factor is at play. In Part II, RNA sequencing analysis and confirmation of select transcriptionally altered operons by RT-qPCR revealed that BpeS might influence expression of the Bsa Type 3 Secretion System (T3SS), while BpeT seems only to target bpeEF-oprC. This may have implication in the pathogenesis of Bp, and must be confirmed in in-vivo cell models using Select Agent excluded strain Bp82 in order to solidify the link between efflux and T3SS during infection. Ultimately, more work is needed to identify the missing regulatory factors in play during expression of bpeEF-oprC, understand how mutations to BpeT and BpeS alter their function, and confirm the relevance of a putative link between co-regulation of efflux and virulence during Bp infection

    Problem Formatting, Domain Specificity, and Arithmetic Processing: The Promise of a Factor Analytic Framework

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    Leading theories of arithmetic cognition take a variety of positions regarding item formatting and its possible effects on encoding, retrieval, and calculation. The extent to which formats might require processing from domains other than mathematics (e.g., a language domain and/or an executive functioning domain) is unclear and an area in need of additional research. The purpose of the current study is to evaluate several leading theories of arithmetic cognition with attention to possible systematic measurement error associated with instrument formatting (method effects) and possible contributions of cognitive domains other than a quantitative domain that is specialized for numeric processing (trait effects). In order to simultaneously examine measurement methods and cognitive abilities, this research is approached from a multi-trait, multi-method factor analytic framework. A sample of 1959 3rd grade students (age M=103.24 months, SD=5.41 months) were selected for the current study from the baseline time points of a larger, longitudinal study conducted in southeastern metropolitan school districts. Abstract Code Theory, Encoding Complex Theory, Triple Code Theory, and the Exact versus Approximate Calculations Hypothesis (a specification of Triple Code Theory) were evaluated with confirmatory factor analysis, using 11 measures of arithmetic with symbolic problem formats (e.g., Arabic numeral and language-based formats) and various problem demands (e.g., requiring both exact and approximate calculations). In general, results from this study provided support for both Triple Code Theory and Encoding Complex Theory, and to some extent, Exact Versus Approximate Calculations Theory is also supported. As predicted by Triple Code Theory, arithmetic outcomes with language formatting, Arabic numeral formatting, and estimation demands across formats were related but distinct from one another. The relationship between problems that required exact calculations (across formats) also provided support for Exact Versus Approximate Calculations Theory’s stipulation that exact calculation problems may draw from the same cognitive processes. As predicted by Encoding Complex Theory, executive function was a direct predictor of all arithmetic outcomes. Language was not a direct predictor of arithmetic outcomes; however, the relationship between language and executive function suggested that language may play a facilitative role in reasoning during numeric processing, particularly for language-formatted problems

    Least Action Principle Applied to a Non-Linear Damped Pendulum

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    The principle of least action is a variational principle that states an object will always take the path of least action as compared to any other conceivable path. This principle can be used to derive the equations of motion of many systems, and therefore provides a unifying equation that has been applied in many fields of physics and mathematics. Hamilton’s formulation of the principle of least action typically only accounts for conservative forces, but can be reformulated to include non-conservative forces such as friction. However, it can be shown that with large values of damping, the object will no longer take the path of least action. Through numerical simulation, this is shown to be true for two simple systems, an object in free fall and a harmonic pendulum, both linearly and cubically damped

    Photograph

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    Photograph

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    Conversations on citizenship: young people\u27s perceptions and performances of democratic citizenship

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    The purpose of this paper is to illuminate how the practice of democratic citizenship compares to ideals of it. I hope to provide a clear view of what contemporary democratic citizenship means, how this conception has been formed, and how democratic citizenship can be effectively practiced to serve the ideals held by democratic citizens. To do this, this paper will be comprised in several parts. After an initial explanation of the theoretical perspective used, I will explore how democracy was understood and implemented in the formative stages of United States government. Next I review contemporary practices of citizenship before examining how presidential primary debates are a reflection of contemporary democracy. Finally, I talk with young people about their perceptions and performances of citizenship. In these conversations I hope to learn how the meaning of citizenship impacts their actions as democratic citizens

    Language Profile and Performances on Math Assessments for Children with Mild Intellectual Disabilities

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    It has been assumed that mathematics testing indicates the development of mathematics concepts, but the linguistic demands of assessment have not been evaluated, especially for children with mild intellectual disabilities. 244 children (grades 2 – 5) were recruited from a larger reading intervention study. Using a multilevel longitudinal SEM model, baseline and post-intervention time points were examined for the contribution of item linguistic complexity, child language skills, and their potential interaction in predicting item level mathematics assessment performance. Item linguistic complexity was an important, stable, and negative predictor of mathematics achievement with children’s language skills significantly and positively predicting mathematics achievement. The interaction between item linguistic complexity and language skills was significant though not stable across time. Following intervention, children with higher language skills performed better on linguistically complex mathematics items. Mathematics achievement may be related to an interaction between children’s language skills and the linguistic demands of the tests themselves

    after “Owls”

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    Monotype

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    Presentation modality influences behavioral measures of alerting, orienting, and executive control

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    The Attention Network Test (ANT) uses visual stimuli to separately assess the attentional skills of alerting (improved performance following a warning cue), spatial orienting (an additional benefit when the warning cue also cues target location), and executive control (impaired performance when a target stimulus contains conflicting information). This study contrasted performance on auditory and visual versions of the ANT to determine whether the measures it obtains are influenced by presentation modality. Forty healthy volunteers completed both auditory and visual tests. Reaction-time measures of executive control were of a similar magnitude and significantly correlated, suggesting that executive control might be a supramodal resource. Measures of alerting were also comparable across tasks. In contrast, spatial-orienting benefits were obtained only in the visual task. Auditory spatial cues did not improve response times to auditory targets presented at the cued location. The different spatial-orienting measures could reflect either separate orienting resources for each perceptual modality, or an interaction between a supramodal orienting resource and modality-specific perceptual processing
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