959 resultados para profluorescent probes


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Soil moisture content, theta, of a bare and vegetated UK gravelly sandy loam soil (in situ and repacked in small lysimeters) was measured using various dielectric instruments (single-sensor ThetaProbes, multi-sensor Profile Probes, and Aquaflex Sensors), at depths ranging between 0.03 and I m, during the summers of 2001 (in situ soil) and 2002 (mini-lysimeters). Half-hourly values of evaporation, E, were calculated from diurnal changes in total soil profile water content, using the soil water balance equation. For the bare soil field, Profile Probes and ML2x ThetaProbes indicated a diurnal course of theta that did not concur with typical soil physical observations: surface layer soil moisture content increased from early morning until about midday, after which theta declined, generally until the early evening. The unexpected course of theta was positively correlated to soil temperature, T-s, also at deeper depths. Aquaflex and ML1 ThetaProbe (older models) outputs, however, reflected common observations: 0 increased slightly during the night (capillary rise) and decreased from the morning until late afternoon (as a result of evaporation). For the vegetated plot, the spurious diurnal theta fluctuations were less obvious, because canopy shading resulted in lower amplitudes of T-s. The unrealistic theta profiles measured for the bare and vegetated field sites caused diurnal estimates of E to attain downward daytime and upward night-time values. In the mini-lysimeters, at medium to high moisture contents, theta values measured by (ML2x) ThetaProbes followed a relatively realistic course, and predictions of E from diurnal changes in vertically integrated theta generally compared well with lysimeter estimates of E. However, time courses of theta and E became comparable to those observed for the field plots when the soil in the lysimeters reached relatively low values of theta. Attempts to correct measured theta for fluctuations in T, revealed that no generally applicable formula could be derived. (c) 2005 Elsevier B.V. All rights reserved.

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The goal of this work is the numerical realization of the probe method suggested by Ikehata for the detection of an obstacle D in inverse scattering. The main idea of the method is to use probes in the form of point source (., z) with source point z to define an indicator function (I) over cap (z) which can be reconstructed from Cauchy data or far. eld data. The indicator function boolean AND (I) over cap (z) can be shown to blow off when the source point z tends to the boundary aD, and this behavior can be used to find D. To study the feasibility of the probe method we will use two equivalent formulations of the indicator function. We will carry out the numerical realization of the functional and show reconstructions of a sound-soft obstacle.

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Jupiter’s magnetosphere acts as a point source of near-relativistic electrons within the heliosphere. In this study, three solar cycles of Jovian electron data in near-Earth space are examined. Jovian electron intensity is found to peak for an ideal Parker spiral connection, but with considerable spread about this point. Assuming the peak in Jovian electron counts indicates the best magnetic connection to Jupiter, we find a clear trend for fast and slow solar wind to be over- and under-wound with respect to the ideal Parker spiral, respectively. This is shown to be well explained in terms of solar wind stream interactions. Thus, modulation of Jovian electrons by corotating interaction regions (CIRs) may primarily be the result of changing magnetic connection, rather than CIRs acting as barriers to cross-field diffusion. By using Jovian electrons to remote sensing magnetic connectivity with Jupiter’s magnetosphere, we suggest that they provide a means to validate solar wind models between 1 and 5 AU, even when suitable in situ solar wind observations are not available. Furthermore, using Jovian electron observations as probes of heliospheric magnetic topology could provide insight into heliospheric magnetic field braiding and turbulence, as well as any systematic under-winding of the heliospheric magnetic field relative to the Parker spiral from footpoint motion of the magnetic field.

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Background: Hexaploid wheat is one of the most important cereal crops for human nutrition. Molecular understanding of the biology of the developing grain will assist the improvement of yield and quality traits for different environments. High quality transcriptomics is a powerful method to increase this understanding. Results: The transcriptome of developing caryopses from hexaploid wheat ( Triticum aestivum, cv. Hereward) was determined using Affymetrix wheat GeneChip (R) oligonucleotide arrays which have probes for 55,052 transcripts. Of these, 14,550 showed significant differential regulation in the period between 6 and 42 days after anthesis ( daa). Large changes in transcript abundance were observed which were categorised into distinct phases of differentiation ( 6 - 10 daa), grain fill ( 12 - 21 daa) and desiccation/maturation ( 28 - 42 daa) and were associated with specific tissues and processes. A similar experiment on developing caryopses grown with dry and/or hot environmental treatments was also analysed, using the profiles established in the first experiment to show that most environmental treatment effects on transcription were due to acceleration of development, but that a few transcripts were specifically affected. Transcript abundance profiles in both experiments for nine selected known and putative wheat transcription factors were independently confirmed by real time RT-PCR. These expression profiles confirm or extend our knowledge of the roles of the known transcription factors and suggest roles for the unknown ones. Conclusion: This transcriptome data will provide a valuable resource for molecular studies on wheat grain. It has been demonstrated how it can be used to distinguish general developmental shifts from specific effects of treatments on gene expression and to diagnose the probable tissue specificity and role of transcription factors.

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The distribution and activity of communities of sulfate-reducing bacteria (SRB) and methanogenic archaea in two contrasting Antarctic sediments were investigated. Methanogenesis dominated in freshwater Lake Heywood, while sulfate reduction dominated in marine Shallow Bay. Slurry experiments indicated that 90% of the methanogenesis in Lake Heywood was acetoclastic. This finding was supported by the limited diversity of clones detected in a Lake Heywood archaeal clone library, in which most clones were closely related to the obligate acetate-utilizing Methanosaeta concilii. The Shallow Bay archaeal clone library contained clones related to the C-1-utilizing Methanolobus and Methanococcoides and the H-2-utilizing Methanogenium. Oligonucleotide probing of RNA extracted directly from sediment indicated that archaea represented 34% of the total prokaryotic signal in Lake Heywood and that Methanosaeta was a major component (13.2%) of this signal. Archaea represented only 0.2% of the total prokaryotic signal in RNA extracted from Shallow Bay sediments. In the Shallow Bay bacterial clone library, 10.3% of the clones were SRB-like, related to Desulfotalea/Desulforhopalus, Desulfofaba, Desulfosarcina, and Desulfobacter as well as to the sulfur and metal oxidizers comprising the Desulfuromonas cluster. Oligonucleotide probes for specific SRB clusters indicated that SRB represented 14.7% of the total prokaryotic signal, with Desulfotalea/Desulforhopalus being the dominant SRB group (10.7% of the total prokaryotic signal) in the Shallow Bay sediments; these results support previous results obtained for Arctic sediments. Methanosaeta and Desulfotalea/Desulforhopalus appear to be important in Lake Heywood and Shallow Bay, respectively, and may be globally important in permanently low-temperature sediments.

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Prebiotics are nondigestible food ingredients that encourage proliferation of selected groups of the colonic microflora, thereby altering the composition toward a more beneficial community. In the present study, the prebiotic potential of a novel galactooligosaccharide (GOS) mixture, produced by the activity of galactosyltransferases from Bifidobacterium bifidum 41171 on lactose, was assessed in vitro and in a parallel continuous randomized pig trial. In situ fluorescent hybridization with 16S rRNA-targeted probes was used to investigate changes in total bacteria, bifidobacteria, lactobacilli, bacteroides, and Clostridium histolyticum group in response to supplementing the novel GOS mixture. In a 3-stage continuous culture system, the bifidobacterial numbers for the first 2 vessels, which represented the proximal and traverse colon, increased (P < 0.05) after the addition of the oligosaccharide mixture. In addition, the oligosaccharide mixture strongly inhibited the attachment of enterohepatic Escherichia coli (P < 0.01) and Salmonella enterica serotype Typhimurium (P < 0.01) to HT29 cells. Addition of the novel mixture at 4% (wt:wt) to a commercial diet increased the density of bificlobacteria (P < 0.001) and the acetate concentration (P < 0.001), and decreased the pH (P < 0.001) compared with the control diet and the control diet supplemented with inulin, suggesting a great prebiotic potential for the novel oligosaccharide mixture. J. Nutr. 135: 1726-1731, 2005.

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Aim: The aim of this study was to measure the gastrointestinal survival of Lactobacillus casei and its impact on the gut microflora in healthy human volunteers. Methods and Results: Twenty healthy volunteers took part in a double-blind placebo-controlled probiotic feeding study (10 fed probiotic, 10 fed placebo). The probiotic was delivered in two 65 ml aliquots of fermented milk drink (FMD) daily for 21 days at a dose of 8.6 +/- 0.1 Log(10)Lact. casei CFU ml(-1) FMD. Faecal samples were collected before, during and after FMD or placebo consumption, and important groups of faecal bacteria enumerated by fluorescent in situ hybridization (FISH) using oligonucleotide probes targeting the 16S rRNA. The fed Lact. casei was enumerated using selective nutrient agar and colony identity confirmed by pulsed field gel electrophoresis. Seven days after ingestion of FMD, the Lact. casei was recovered from faecal samples taken from the active treatment group at 7.1 +/- 0.4 Log(10) CFU g(-1) faeces (mean +/- SD, n = 9) and numbers were maintained at this level until day 21. Lact. casei persisted in six volunteers until day 28 at 5.0 +/- 0.9 Log(10) CFU g(-1) faeces (mean +/- SD, n = 6). Numbers of faecal lactobacilli increased significantly upon FMD ingestion. In addition, the numbers of bifidobacteria were higher on days 7 and 21 than on days 0 and 28 in both FMD fed and placebo fed groups. Consumption of Lact. casei had little discernible effect on other bacterial groups enumerated. Conclusions: Daily consumption of FMD enabled a probiotic Lact. casei strain to be maintained in the gastrointestinal tract of volunteers at a stable relatively high population level during the probiotic feeding period. Significance and Impact of the Study: The study has confirmed that this probiotic version of Lact. casei survives well within the human gastrointestinal tract.

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Exopolysaccharides (EPS) isolated from two Bifidobacterium strains, one of human intestinal origin (Bifidobacterium longum subsp. longum IPLA E44) and the other from dairy origin (Bifidobacterium animalis subsp. lactis IPLA R1), were subjected to in vitro chemically simulated gastrointestinal digestion. which showed the absence of degradation of both polymers in these conditions. Polymers were then used as carbon sources in pH-controlled faecal batch cultures and compared with the non-prebiotic carbohydrate glucose and the prebiotic inulin to determine changes in the composition of faecal bacteria. A set of eight fluorescent in situ hybridisation oligonucleotide probes targeting 16S rRNA sequences was used to quantify specific groups of microorganisms. Growth of the opportunistic pathogen Clostridium histolyticum occurred with all carbohydrates tested similarly to that found in negative control cultures without added carbohydrate and was mainly attributed to the culture conditions used rather than enhancement of growth by these substrates. Polymers E44 and RI stimulated growth of Lactobacillus/Enterococcus, Bifidobacterium, and Bacteroides/Prevotella in a similar way to that seen with inulin. The EPS RI also promoted growth of the Atopobium cluster during the first 24 h of fermentation. An increase in acetic and lactic acids was found during early stages of fermentation (first 10-24 h) correlating with increases of Lactobacillus, Bifidobacterium, and Atopobium. Propionic acid concentrations increased in old cultures, which was coincident with the enrichment of Clostridium cluster IX in cultures with EPS RI and with the increases in Bacteroides in cultures with both microbial EPS (RI and E44) and inulin. The lowest acetic to propionic acid ratio was obtained for EPS E44. None of the carbohydrates tested supported the growth of microorganisms from Clostridium clusters XIVa+b and IV, results that correlate with the poor butyrate production in the presence of EPS. Thus, EPS synthesized by bifidobacteria from dairy and intestinal origins can modulate the intestinal microbiota in vitro, promoting changes in some numerically and metabolically relevant microbial populations and shifts in the production of short chain fatty acids. (C) 2009 Elsevier B.V. All rights reserved.

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Children with autistic spectrum disorders (ASDs) tend to suffer from severe gastrointestinal problems. Such symptoms may be due to a disruption of the indigenous gut flora promoting the overgrowth of potentially pathogenic micro-organisms. The faecal flora of patients with ASDs was studied and compared with those of two control groups (healthy siblings and unrelated healthy children). Faecal bacterial populations were assessed through the use of a culture-independent technique, fluorescence in situ hybridization, using oligonucleotide probes targeting predominant components of the gut flora. The faecal flora of ASD patients contained a higher incidence of the Clostridium histolyticum group (Clostridium clusters I and 11) of bacteria than that of healthy children. However, the non-autistic sibling group had an intermediate level of the C. histolyticum group, which was not significantly different from either of the other subject groups. Members of the C. histolyticum group are recognized toxin-producers and may contribute towards gut dysfunction, with their metabolic products also exerting systemic effects. Strategies to reduce clostridial population levels harboured by ASD patients or to improve their gut microflora profile through dietary modulation may help to alleviate gut disorders common in such patients.

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The antioxidant activity of hydroxytyrosol, hydroxytyrosol acetate, oleuropein, 3,4-dihydroxyphenylelenolic acid (3,4-DHPEA-EA) and 3,4-dihydroxyphenyielenolic acid dialdehyde (3,4-DHPEA-EDA) towards oxidation initiated by 2,2'-azobis (2-amidinopropane) hydrochloride in a soybean phospholipid liposome system was studied. The antioxidant activity of these olive oil phenols was similar and the duration of the lag phase was almost twice that of alpha-tocopherol. Trolox(R), a water-soluble analogue of alpha-tocopherol, showed the worst antioxidant activity. However, oxidation before the end of the lag phase was inhibited less effectively by the olive oil phenols than by alpha-tocopherol and Trolox(R). Synergistic effects (11-20% increase in lag phase) were observed in the antioxidant activity of combinations of alpha-tocopherol with olive oil phenols both with and without ascorbic acid. Fluorescence anisotropy of probes and fluorescence quenching studies showed that the olive oil phenols did not penetrate into the membrane, but their effectiveness as antioxidants showed they were associated with the surface of the phospholipid bilayer. (C) 2003 Elsevier Science Ireland Ltd. All rights reserved.

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It is evident that quantitative information on different microbial groups and their contribution in terms of activity in the gastrointestinal (GI) tract of humans and animals is required in order to formulate functional diets targeting improved gut function and host health. In this work, quantitative information on levels and spatial distributions of Bacteroides spp, Eubacterium spp, Clostridium spp, Escherichia coli, Bifidobacterium spp and Lactobacillus/Enterococcus spp. along the porcine large intestine was investigated using 16S rRNA targeted probes and fluorescent in situ hybridisation (FISH). Caecum, ascending colon (AC) and rectum luminal digesta from three groups of individually housed growing pigs fed either a corn-soybean basal diet (CON diet) or a prebiotic diet containing 10 g/kg oligofructose (FOS diet) or trans-galactooligosaccharides (TOS diet) at the expense of cornstarch were analysed. DAPI staining was used to enumerate total number of cells in the samples. Populations of total cells, Bacteroides, Eubacterium, Clostridium and Bifidobacterium, declined significantly (P < 0.05) from caecum to rectum, and were not affected by dietary treatments. Populations of Lactobacillus/ Enterococcus and E coli did not differ throughout the large intestine. The relative percent (%) contribution of each bacterial group to the total cell count did not differ between caecum and rectum, with the exception of Eubacterium that was higher in the AC digesta. FISH analysis showed that the sum of all bacterial groups made up a small percentage of the total cells, which was 12.4%, 21.8% and 10.3% in caecum, AC and rectum, respectively. This supports the view that in swine, the diversity of GI microflora might be higher compared to other species. In terms of microflora metabolic activity, the substantially higher numerical trends seen in FOS and TOS treatments regarding total volatile fatty acid, acetate concentrations and glycolytic activities, it could be postulated that FOS and TOS promoted saccharolytic activities in the porcine colon. (c) 2006 Elsevier Ltd. All rights reserved.

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The composition of the colonic microbiota of 91 northern Europeans was characterized by fluorescent in situ hybridization using 18 phylogenetic probes. On average 75% of the bacteria were identified, and large interindividual variations were observed. Clostridium coccoides and Clostridium leptum were the dominant groups (28.0% and 25.2%), followed by the Bacteroides (8.5%). According to principal component analysis, no significant grouping with respect to geographic origin, age, or gender was observed.

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Recent large-scale cloning studies have shown that the ratio of Bacteroidetes to Firmicutes may be important in the obesity-associated gut microbiota, but the species these phyla represent in this ecosystem has not been examined. The Bacteroidetes data from the recent Turnbaugh study were examined to determine those members of the phylum detected in human faecal samples. In addition, FISH analysis was performed on faecal samples from 17 healthy, nonobese donors using probe Bac303, routinely used by gut microbiologists to enumerate BacteroidesPrevotella populations in faecal samples, and another probe (CFB286) whose target range has some overlap with that of Bac303. Sequence analysis of the Turnbaugh data showed that 23/519 clones were chimeras or erroneous sequences; all good sequences were related to species of the order Bacteroidales, but no one species was present in all donors. FISH analysis demonstrated that approximately one-quarter of the healthy, nonobese donors harboured high numbers of Bacteroidales not detected by probe Bac303. It is clear that Bacteroidales populations in human faecal samples have been underestimated in FISH-based studies. New probes and complementary primer sets should be designed to examine numerical and compositional changes in the Bacteroidales during dietary interventions and in studies of the obesity-associated microbiota in humans and animal model systems.

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The Maillard reaction causes changes to protein structure and occurs in foods mainly during thermal treatment. Melanoidins, the final products of the Maillard reaction, may enter the gastrointestinal tract, which is populated by different species of bacteria. In this study, melanoidins were prepared from gluten and glucose. Their effect on the growth of faecal bacteria was determined in culture with genotype and phenotype probes to identify the different species involved. Analysis of peptic and tryptic digests showed that low molecular mass products are formed from the degradation of melanoidins. Results showed a change in the growth of bacteria. This in vitro study demonstrated that melanoidins, prepared from gluten and glucose, affect the growth of the gut microflora.

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High spatial resolution vertical profiles of pore-water chemistry have been obtained for a peatland using diffusive equilibrium in thin films (DET) gel probes. Comparison of DET pore-water data with more traditional depth-specific sampling shows good agreement and the DET profiling method is less invasive and less likely to induce mixing of pore-waters. Chloride mass balances as water tables fell in the early summer indicate that evaporative concentration dominates and there is negligible lateral flow in the peat. Lack of lateral flow allows element budgets for the same site at different times to be compared. The high spatial resolution of sampling also enables gradients to be observed that permit calculations of vertical fluxes. Sulfate concentrations fall at two sites with net rates of 1.5 and 5.0nmol cm− 3 day− 1, likely due to a dominance of bacterial sulfate reduction, while a third site showed a net gain in sulfate due to oxidation of sulfur over the study period at an average rate of 3.4nmol cm− 3 day− 1. Behaviour of iron is closely coupled to that of sulfur; there is net removal of iron at the two sites where sulfate reduction dominates and addition of iron where oxidation dominates. The profiles demonstrate that, in addition to strong vertical redox related chemical changes, there is significant spatial heterogeneity. Whilst overall there is evidence for net reduction of sulfate within the peatland pore-waters, this can be reversed, at least temporarily, during periods of drought when sulfide oxidation with resulting acid production predominates.