162 research outputs found

    A Rapid Subtractive Immunization Method to Prepare Discriminatory Monoclonal Antibodies for Food E. coli O157:H7 Contamination

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    To detect food E. coli O157:H7 contamination rapidly and accurately, it is essential to prepare high specific monoclonal antibodies (mAbs) against the pathogen. Cyclophosphamide (Cy)-mediated subtractive immunization strategy was performed in mice to generate mAbs that react with E. coli O157:H7, but not with other affiliated bacteria. Specificity of 19 mAbs was evaluated by ELISA and/or dot-immunogold filtration assay (DIGFA). Immunogloubin typing, affinity and binding antigens of 5 selected mAbs were also analysed. MAbs 1D8, 4A7, 5A2 were found to have high reactivity with E. coli O157:H7 and no cross-reactivity with 80 other strains of bacteria including Salmonella sp., Shigella sp., Proteus sp., Yersinia enterocolitica, Staphylococcus aureus, Klebsiella pneumoniae, Citrobacter freundii and other non-E. coli O157:H7 enteric bacteria. Their ascetic titers reached 1∶106 with E. coli O157:H7 and affinity constants ranged from 1.57×1010 to 2.79×1010 L/mol. The antigens recognized by them were different localized proteins. Furthermore, immune-colloidal gold probe coated with mAb 5A2 could specifically distinguish minced beef contaminated by E. coli O157:H7 from 84 other bacterial contaminations. The Cy-mediated subtractive immunization procedure coupled with hybridoma technology is a rapid and efficient approach to prepare discriminatory mAbs for detection of E. coli O157:H7 contamination in food

    Fluorescent RNA cytosine analogue - an internal probe for detailed structure and dynamics investigations

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    The bright fluorescent cytosine analogue tCO stands out among fluorescent bases due to its virtually unquenched fluorescence emission in duplex DNA. However, like most reported base analogues, it has not been thoroughly characterized in RNA. We here report on the first synthesis and RNA-incorporation of tCO, and characterize its base-mimicking and fluorescence properties in RNA. As in DNA, we find a high quantum yield inside RNA duplexes (<?F> = 0.22) that is virtually unaffected by the neighbouring bases (?F = 0.20-0.25), resulting in an average brightness of 1900 M-1 cm-1. The average fluorescence lifetime in RNA duplexes is 4.3 ns and generally two lifetimes are required to fit the exponential decays. Fluorescence properties in ssRNA are defined by a small increase in average quantum yield (<?F > = 0.24) compared to dsRNA, with a broader distribution (?F = 0.17-0.34) and slightly shorter average lifetimes. Using circular dichroism, we find that the tCO-modified RNA duplexes form regular A-form helices and in UV-melting experiments the stability of the duplexes is only slightly higher than that of the corresponding natural RNA (<?T m> = + 2.3 °C). These properties make tCO a highly interesting fluorescent RNA base analogue for detailed FRET-based structural measurements, as a bright internal label in microscopy, and for fluorescence anisotropy measurements of RNA dynamics

    Is diagnosing exposed dentine a suitable tool for grading erosive loss?

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    Quantifying tooth wear in general and erosion in particular mostly is made by distinguishing between lesions restricted to enamel and lesions reaching the underlying dentine. Various scores for grading have been used, but in all systems, higher scores are given in cases of exposed dentine, thus, indicating a more severe stage of the condition. Clinical diagnosis of exposed dentine is made by assessing changes in colour or optical properties of the hard tissues. This paper aims to review the literature and discuss critically problems arising form this approach. It appears that classifying the severity of erosion by the area or depth of exposed dentine is difficult and poorly reproducible, and taking into account the variation of enamel thickness, the amount of tissue lost often is not related simply to the area of exposed dentine. There has still been very little longitudinal investigation of the significance of exposed dentine as a prognostic indicator. Further work and discussion is needed to reevaluate the explanative power of current grading procedures

    High-quality habitat and facilitation ameliorate competitive effects of prior residents on new settlers

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    Many species disperse during their lifetime. Two factors that can affect the performance of individuals following dispersal are the presence of conspecifics and intrinsic habitat quality at the settlement site. Detecting the influence of these factors can be difficult for at least two reasons: (1) the outcomes of interactions with conspecifics are often variable including both competition and facilitation, and (2) selection of high quality habitats often leads to positive covariance between habitat quality and density. In this study, I investigate positive and negative effects of resident blue streak cleaner wrasse (Labroides dimidiatus) on the growth and survival of recently settled conspecifics while accounting for habitat quality. Juvenile L. dimidiatus settle near adult conspecifics, but likely have to compete with resident adults for access to food. However, field experiments indicate that settlers have access to more resources at occupied sites, and as a result, grow faster despite evidence for competition with residents. This result is a direct consequence of two factors: (1) resident conspecifics facilitate settlers by attracting client fish, and (2) resident conspecifics are strongly associated with high quality habitat. These results highlight the need to simultaneously consider habitat quality and competitive and facilitative interactions between conspecifics when making inferences about ecological processes from spatial patterns of individual performance

    Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

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    In order to establish infections within the mammalian host, pathogens must protect themselves against toxic reactive oxygen species produced by phagocytes of the immune system. The fungal pathogen Histoplasma capsulatum infects both neutrophils and macrophages but the mechanisms enabling Histoplasma yeasts to survive in these phagocytes have not been fully elucidated. We show that Histoplasma yeasts produce a superoxide dismutase (Sod3) and direct it to the extracellular environment via N-terminal and C-terminal signals which promote its secretion and association with the yeast cell surface. This localization permits Sod3 to protect yeasts specifically from exogenous superoxide whereas amelioration of endogenous reactive oxygen depends on intracellular dismutases such as Sod1. While infection of resting macrophages by Histoplasma does not stimulate the phagocyte oxidative burst, interaction with polymorphonuclear leukocytes (PMNs) and cytokine-activated macrophages triggers production of reactive oxygen species (ROS). Histoplasma yeasts producing Sod3 survive co-incubation with these phagocytes but yeasts lacking Sod3 are rapidly eliminated through oxidative killing similar to the effect of phagocytes on Candida albicans yeasts. The protection provided by Sod3 against host-derived ROS extends in vivo. Without Sod3, Histoplasma yeasts are attenuated in their ability to establish respiratory infections and are rapidly cleared with the onset of adaptive immunity. The virulence of Sod3-deficient yeasts is restored in murine hosts unable to produce superoxide due to loss of the NADPH-oxidase function. These results demonstrate that phagocyte-produced ROS contributes to the immune response to Histoplasma and that Sod3 facilitates Histoplasma pathogenesis by detoxifying host-derived reactive oxygen thereby enabling Histoplasma survival

    Seed Dispersal Anachronisms: Rethinking the Fruits Extinct Megafauna Ate

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    Background: Some neotropical, fleshy-fruited plants have fruits structurally similar to paleotropical fruits dispersed by megafauna (mammals.10 3 kg), yet these dispersers were extinct in South America 10–15 Kyr BP. Anachronic dispersal systems are best explained by interactions with extinct animals and show impaired dispersal resulting in altered seed dispersal dynamics. Methodology/Principal Findings: We introduce an operational definition of megafaunal fruits and perform a comparative analysis of 103 Neotropical fruit species fitting this dispersal mode. We define two megafaunal fruit types based on previous analyses of elephant fruits: fruits 4–10 cm in diameter with up to five large seeds, and fruits.10 cm diameter with numerous small seeds. Megafaunal fruits are well represented in unrelated families such as Sapotaceae, Fabaceae, Solanaceae, Apocynaceae, Malvaceae, Caryocaraceae, and Arecaceae and combine an overbuilt design (large fruit mass and size) with either a single or few (,3 seeds) extremely large seeds or many small seeds (usually.100 seeds). Within-family and within-genus contrasts between megafaunal and non-megafaunal groups of species indicate a marked difference in fruit diameter and fruit mass but less so for individual seed mass, with a significant trend for megafaunal fruits to have larger seeds and seediness. Conclusions/Significance: Megafaunal fruits allow plants to circumvent the trade-off between seed size and dispersal b

    The LUX-ZEPLIN (LZ) Experiment

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    We describe the design and assembly of the LUX-ZEPLIN experiment, a direct detection search for cosmic WIMP dark matter particles. The centerpiece of the experiment is a large liquid xenon time projection chamber sensitive to low energy nuclear recoils. Rejection of backgrounds is enhanced by a Xe skin veto detector and by a liquid scintillator Outer Detector loaded with gadolinium for efficient neutron capture and tagging. LZ is located in the Davis Cavern at the 4850' level of the Sanford Underground Research Facility in Lead, South Dakota, USA. We describe the major subsystems of the experiment and its key design features and requirements
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