Individual-specific changes in the human gut microbiota after challenge with enterotoxigenic Escherichia coli and subsequent ciprofloxacin treatment

IA, JNP, and MP were partly supported by the NIH, grants R01-AI-100947 to MP, and R21-GM-107683 to Matthias Chung, subcontract to MP. JNP was partly supported by an NSF graduate fellowship number DGE750616. IA, JNP, BRL, OCS and MP were supported in part by the Bill and Melinda Gates Foundation, award number 42917 to OCS. JP and AWW received core funding support from The Wellcome Trust (grant number 098051). AWW, and the Rowett Institute of Nutrition and Health, University of Aberdeen, receive core funding support from the Scottish Government Rural and Environmental Science and Analysis Service (RESAS).

Spontaneous mutations recovered as mosaics in the mouse specific-locus test

The specific-locus test (SLT) detects new mutants among mice heterozygous for seven recessive visible markers. Spontaneous mutations can be manifested not only as singleton whole-body mutants in controls (for which we report new data), but as mosaics—either visible (manifesting mottled coat color) in the scored generation (G2) or masked, among the wild-type parental generation (G1). Masked G1 mosaics reveal themselves by producing clusters of whole-body mutants in G2. We provide evidence that most, if not all, mosaics detected in the SLT (both radiation and control progenies) result from a single-strand spontaneous mutation subsequent to the last premeiotic mitosis and before the first postmeiotic one of a parental genome—the “perigametic interval.” Such events in the genomes of the G1 and G0 result, respectively, in visible and masked 50:50 mosaics. Per cell cycle, the spontaneous mutation rate in the perigametic interval is much higher than that in pregamete mitotic divisions. A clearly different locus spectrum further supports the hypothesis of different origin, and casts further doubt on the validity of the doubling-dose risk-estimation method. Because mosaics cannot have arisen in mitotic germ cells, and are not induced by radiation exposure in the perigametic interval, they should not be included in calculations of radiation-induced germ-line mutation rates. For per-generation calculations, inclusion of mosaics yields a spontaneous frequency 1.7 times that calculated from singletons alone for mutations contributed by males; including both sexes, the multiple is 2.2.

An experimental test for lineage-specific position effects on alcohol dehydrogenase (Adh) genes in Drosophila

Independent transgene insertions differ in expression based on their location in the genome; these position effects are of interest because they reflect the influence of genome organization on gene regulation. Position effects also represent potentially insurmountable obstacles to the rigorous functional comparison of homologous genes from different species because (i) quantitative variation in expression of each gene across genomic positions (generalized position effects, or GPEs) may overwhelm differences between the genes of interest, or (ii) divergent genes may be differentially sensitive to position effects, reflecting unique interactions between each gene and its genomic milieu (lineage-specific position effects, or LSPEs). We have investigated both types of position-effect variation by applying our method of transgene coplacement, which allows comparisons of transgenes in the same position in the genome of Drosophila melanogaster. Here we report an experimental test for LSPE in Drosophila. The alcohol dehydrogenase (Adh) genes of D. melanogaster and Drosophila affinidisjuncta differ in both tissue distribution and amounts of ADH activity. Despite this striking regulatory divergence, we found a very high correlation in overall ADH activity between the genes of the two species when placed in the same genomic position as assayed in otherwise Adh-null adults and larvae. These results argue against the influence of LSPE for these sequences, although the effects of GPE are significant. Our new findings validate the coplacement approach and show that it greatly magnifies the power to detect differences in expression between transgenes. Transgene coplacement thus dramatically extends the range of functional and evolutionary questions that can be addressed by transgenic technology.

A quantitative test to estimate neutralizing antibodies to the hepatitis C virus: cytofluorimetric assessment of envelope glycoprotein 2 binding to target cells.

Hepatitis C virus (HCV) is a major cause of chronic hepatitis. The virus does not replicate efficiently in cell cultures, and it is therefore difficult to assess infection-neutralizing antibodies and to evaluate protective immunity in vitro. To study the binding of the HCV envelope to cell-surface receptors, we developed an assay to assess specific binding of recombinant envelope proteins to human cells and neutralization thereof. HCV recombinant envelope proteins expressed in various systems were incubated with human cells, and binding was assessed by flow cytometry using anti-envelope antibodies. Envelope glycoprotein 2 (E2) expressed in mammalian cells, but not in yeast or insect cells, binds human cells with high affinity (Kd approximately 10(-8) M). We then assessed antibodies able to neutralize E2 binding in the sera of both vaccinated and carrier chimpanzees, as well as in the sera of humans infected with various HCV genotypes. Vaccination with recombinant envelope proteins expressed in mammalian cells elicited high titers of neutralizing antibodies that correlated with protection from HCV challenge. HCV infection does not elicit neutralizing antibodies in most chimpanzees and humans, although low titers of neutralizing antibodies were detectable in a minority of infections. The ability to neutralize binding of E2 derived from the HCV-1 genotype was equally distributed among sera from patients infected with HCV genotypes 1, 2, and 3, demonstrating that binding of E2 is partly independent of E2 hypervariable regions. However, a mouse monoclonal antibody raised against the E2 hypervariable region 1 can partially neutralize binding of E2, indicating that at least two neutralizing epitopes, one of which is hypervariable, should exist on the E2 protein. The neutralization-of-binding assay described will be useful to study protective immunity to HCV infection and for vaccine development.

Days of challenge, years of change : a technical history of the Pacific Missile Test Center.

Includes index.

Rational drawdown test of artesian well elucidates specific capacity /

Mode of access: Internet.

Evaluating the properties of a stage-specific self-efficacy scale for physical activity using classical test theory, confirmatory factor analysis and item response modeling

The purpose of this paper was to evaluate the psychometric properties of a stage-specific selfefficacy scale for physical activity with classical test theory (CTT), confirmatory factor analysis (CFA) and item response modeling (IRM). Women who enrolled in the Women On The Move study completed a 20-item stage-specific self-efficacy scale developed for this study [n = 226, 51.1% African-American and 48.9% Hispanic women, mean age = 49.2 (67.0) years, mean body mass index = 29.7 (66.4)]. Three analyses were conducted: (i) a CTT item analysis, (ii) a CFA to validate the factor structure and (iii) an IRM analysis. The CTT item analysis and the CFA results showed that the scale had high internal consistency (ranging from 0.76 to 0.93) and a strong factor structure. Results also showed that the scale could be improved by modifying or eliminating some of the existing items without significantly altering the content of the scale. The IRM results also showed that the scale had few items that targeted high self-efficacy and the stage-specific assumption underlying the scale was rejected. In addition, the IRM analyses found that the five-point response format functioned more like a four-point response format. Overall, employing multiple methods to assess the psychometric properties of the stage-specific self-efficacy scale demonstrated the complimentary nature of these methods and it highlighted the strengths and weaknesses of this scale.

Development of a co-culture model of the human lungs for toxicity testing and identification of biomarkers of inhalation toxicity

The airway epithelium is the first point of contact in the lung for inhaled material, including infectious pathogens and particulate matter, and protects against toxicity from these substances by trapping and clearance via the mucociliary escalator, presence of a protective barrier with tight junctions and initiation of a local inflammatory response. The inflammatory response involves recruitment of phagocytic cells to neutralise and remove and invading materials and is oftern modelled using rodents. However, development of valid in vitro airway epithelial models is of great importance due to the restrictions on animal studies for cosmetic compound testing implicit in the 7th amendment to the European Union Cosmetics Directive. Further, rodent innate immune responses have fundamental differences to human. Pulmonary endothelial cells and leukocytes are also involved in the innate response initiated during pulmonary inflammation. Co-culture models of the airways, in particular where epithelial cells are cultured at air liquid interface with the presence of tight junctions and differentiated mucociliary cells, offer a solution to this problem. Ideally validated models will allow for detection of early biomarkers of response to exposure and investigation into inflammatory response during exposure. This thesis describes the approaches taken towards developing an in vitro epithelial/endothelial cell model of the human airways and identification biomarkers of response to exposure to xenobiotics. The model comprised normal human primary microvascular endothelial cells and the bronchial epithelial cell line BEAS-2B or normal human bronchial epithelial cells. BEAS-2B were chosen as their characterisation at air liquid interface is limited but they are robust in culture, thereby predicted to provide a more reliable test system. Proteomics analysis was undertaken on challenged cells to investigate biomarkers of exposure. BEAS-2B morphology was characterised at air liquid interface compared with normal human bronchial epithelial cells. The results indicate that BEAS-2B cells at an air liquid interface form tight junctions as shown by expression of the tight junction protein zonula occludens-1. To this author’s knowledge this is the first time this result has been reported. The inflammatory response of BEAS-2B (measured as secretion of the inflammatory mediators interleukin-8 and -6) air liquid interface mono-cultures to Escherichia coli lipopolysaccharide or particulate matter (fine and ultrafine titanium dioxide) was comparable to published data for epithelial cells. Cells were also exposed to polymers of “commercial interest” which were in the nanoparticle range (and referred to particles hereafter). BEAS-2B mono-cultures showed an increased secretion of inflammatory mediators after challenge. Inclusion of microvascular endothelial cells resulted in protection against LPS- and particle- induced epithelial toxicity, measured as cell viability and inflammatory response, indicating the importance of co-cultures for investigations into toxicity. Two-dimensional proteomic analysis of lysates from particle-challenged cells failed to identify biomarkers of toxicity due to assay interference and experimental variability. Separately, decreased plasma concentrations of serine protease inhibitors, and the negative acute phase proteins transthyretin, histidine-rich glycoprotein and alpha2-HS glycoprotein were identified as potential biomarkers of methyl methacrylate/ethyl methacrylate/butylacrylate treatment in rats.

Enhancing the Manufacturing Knowledge of Undergraduate Engineering Students: A Case Study of a Design-Build-Test Challenge Involving Folding Bicycles

Many engineers currently in professional practice will have gained a degree level qualification which involved studying a curriculum heavy with mathematics and engineering science. While this knowledge is vital to the engineering design process so also is manufacturing knowledge, if the resulting designs are to be both technically and commercially viable.
The methodology advanced by the CDIO Initiative aims to improve engineering education by teaching in the context of Conceiving, Designing, Implementing and Operating products, processes or systems. A key element of this approach is the use of Design-Built-Test (DBT) projects as the core of an integrated curriculum. This approach facilitates the development of professional skills as well as the application of technical knowledge and skills developed in other parts of the degree programme. This approach also changes the role of lecturer to that of facilitator / coach in an active learning environment in which students gain concrete experiences that support their development.
The case study herein describes Mechanical Engineering undergraduate student involvement in the manufacture and assembly of concept and functional prototypes of a folding bicycle.

The Geometric Curvature Of The Spine Of Runners During Maximal Incremental Effort Test.

This study sought to analyse the behaviour of the average spinal posture using a novel investigative procedure in a maximal incremental effort test performed on a treadmill. Spine motion was collected via stereo-photogrammetric analysis in thirteen amateur athletes. At each time percentage of the gait cycle, the reconstructed spine points were projected onto the sagittal and frontal planes of the trunk. On each plane, a polynomial was fitted to the data, and the two-dimensional geometric curvature along the longitudinal axis of the trunk was calculated to quantify the geometric shape of the spine. The average posture presented at the gait cycle defined the spine Neutral Curve. This method enabled the lateral deviations, lordosis, and kyphosis of the spine to be quantified noninvasively and in detail. The similarity between each two volunteers was a maximum of 19% on the sagittal plane and 13% on the frontal (p<0.01). The data collected in this study can be considered preliminary evidence that there are subject-specific characteristics in spinal curvatures during running. Changes induced by increases in speed were not sufficient for the Neutral Curve to lose its individual characteristics, instead behaving like a postural signature. The data showed the descriptive capability of a new method to analyse spinal postures during locomotion; however, additional studies, and with larger sample sizes, are necessary for extracting more general information from this novel methodology.

Da avaliação a gestão de processo : uma proposta de instrumento para acompanhamento da inclusão contextualizada no transcorrer de atividades motoras

Universidade Estadual de Campinas . Faculdade de Educação Física

Sensitization Prevalence, Antibody Cross-Reactivity and Immunogenic Peptide Profile of Api g 2, the Non-Specific Lipid Transfer Protein 1 of Celery

Background: Celery (Apium graveolens) represents a relevant allergen source that can elicit severe reactions in the adult population. To investigate the sensitization prevalence and cross-reactivity of Api g 2 from celery stalks in a Mediterranean population and in a mouse model. Methodology: 786 non-randomized subjects from Italy were screened for IgE reactivity to rApi g 2, rArt v 3 (mugwort pollen LTP) and nPru p 3 (peach LTP) using an allergen microarray. Clinical data of 32 selected patients with reactivity to LTP under investigation were evaluated. Specific IgE titers and cross-inhibitions were performed in ELISA and allergen microarray. Balb/c mice were immunized with purified LTPs; IgG titers were determined in ELISA and mediator release was examined using RBL-2H3 cells. Simulated endolysosomal digestion was performed using microsomes obtained from human DCs. Results: IgE testing showed a sensitization prevalence of 25.6% to Api g 2, 18.6% to Art v 3, and 28.6% to Pru p 3 and frequent co-sensitization and correlating IgE-reactivity was observed. 10/32 patients suffering from LTP-related allergy reported symptoms upon consumption of celery stalks which mainly presented as OAS. Considerable IgE cross-reactivity was observed between Api g 2, Art v 3, and Pru p 3 with varying inhibition degrees of individual patients' sera. Simulating LTP mono-sensitization in a mouse model showed development of more congruent antibody specificities between Api g 2 and Art v 3. Notably, biologically relevant murine IgE cross-reactivity was restricted to the latter and diverse from Pru p 3 epitopes. Endolysosomal processing of LTP showed generation of similar clusters, which presumably represent T-cell peptides. Conclusions: Api g 2 represents a relevant celery stalk allergen in the LTP-sensitized population. The molecule displays common B cell epitopes and endolysosomal peptides that encompass T cell epitopes with pollen and plant-food derived LTP.

A special judo fitness test classificatory table

Background and Study Aim: Judo is very physiological demanding sport, but there are no many physical fitness specific tests. One of the most used specific judo tests is the Special Judo Fitness Test (SJFT) proposed by Sterkowicz ( 1995). Although this test has been used by many coaches in different countries no classificatory table was found to classify the judo athletes according to their results. Thus, the aim of this work was to present a classificatory table for this test. Material/Methods: For this purpose 141 judo athletes ( mean +/-standard deviation: 21.3+/-4.5years-old; 74.2+/-15.9 kg of body mass and 176.7+/-8.2 cm of height; judo ranking between 3(rd) kyu and 3(rd) dan) familiarized with the SJFT performed it once in order to provide data to establish a classificatory table. Results: After the analysis of data distribution a five scale table (20% for each classificatory category) was developed considering the variables used in the SJFT ( number of throws, heart rate after and 1 min after the test and index). Conclusions: The classificatory table can help coaches using the SJFT to classify their athletes` level and to monitor their physical fitness progress.

A Semi-Tethered Test for Power Assessment in Running

We analyzed the usefulness of a semi-tethered field running test (STR) and the relationships between indices of anaerobic power, anaerobic capacity and running performance in 9 trained male sprinters (22.2 +/- 2.9 yrs, 176 +/- 1 cm, 68.0 +/- 9.4 kg). STR involved an all out 120 m run attached to an apparatus that enabled power calculation from force and velocity measures. Subjects also carried out a cycloergometer Win-gate Anaerobic Test (WT), an all out 300 m run and had accessed their maximal accumulated oxygen deficit (MAOD) on a treadmill. Peak and mean powers attained in STR (1 720 +/- 221 and 1 391 +/- 201 W) were greater but significantly related (r=0.82; P<0.01) to those in the WT (808 +/- 130 and 603 +/- 87 W). In addition, power measures derived from the STR were stronger related to running performance compared to those from the WT (r=0.81-0.94 vs. 0.68-0.84; P<0.05). Relationships between MAOD and most power indices were only weak to moderate. These results support the usefulness of STR for specific power assessment in field running and suggest that anaerobic power and capacity are not related entities, irrespective of having been evaluated using similar or dissimilar exercise modes.