Evaluation of the environmental specificity of fluorescence in situ hybridization (FISH) using fluorescence-activated cell sorting (FACS) of probe (PSE1284)-positive cells extracted from rhizosphere soil

We explicitly tested for the first time the ‘environmental specificity’ of traditional 16S rRNAtargeted fluorescence in situ hybridization (FISH) through comparison of the bacterial diversity actually targeted in the environment with the diversity that should be exactly targeted (i.e. without mismatches) according to in silico analysis. To do this, we exploited advances in modern Flow Cytometry that enabled improved detection and therefore sorting of sub-micron-sized particles and used probe PSE1284 (designed to target Pseudomonads) applied to Lolium perenne rhizosphere soil as our test system. The 6-carboxyfluorescein (6-FAM)-PSE1284-hybridised population, defined as displaying enhanced green fluorescence in Flow Cytometry, represented 3.51±1.28% of the total detected population when corrected using a nonsense (NON-EUB338) probe control. Analysis of 16S rRNA gene libraries constructed from Fluorescence Activated Cell Sorted (FACS) -recovered fluorescent populations (n=3), revealed that 98.5% (Pseudomonas spp. comprised 68.7% and Burkholderia spp. 29.8%) of the total sorted population was specifically targeted as evidenced by the homology of the 16S rRNA sequences to the probe sequence. In silico evaluation of probe PSE1284 with the use of RDP-10 probeMatch justified the existence of Burkholderia spp. among the sorted cells. The lack of novelty in Pseudomonas spp. sequences uncovered was notable, probably reflecting the well-studied nature of this functionally important genus. To judge the diversity recorded within the FACS-sorted population, rarefaction and DGGE analysis were used to evaluate, respectively, the proportion of Pseudomonas diversity uncovered by the sequencing effort and the representativeness of the Nycodenz® method for the extraction of bacterial cells from soil.

In situ preservation of wetland heritage: hydrological and chemical change in the burial environment of the Somerset Levels, UK.

In Situ preservation is a core strategy for the conservation and management of waterlogged remains at wetland sites. Inorganic and organic remains can, however, quickly become degraded, or lost entirely, as a result of chemical or hydrological changes. Monitoring is therefore crucial in identifying baseline data for a site, the extent of spatial and or temporal variability, and in evaluating the potential impacts of these variables on current and future In Situ preservation potential. Since August 2009, monthly monitoring has taken place at the internationally important Iron Age site of Glastonbury Lake Village in the Somerset Levels, UK. A spatial, stratigraphic, and analytical approach to the analysis of sediment horizons and monitoring of groundwater chemistry, redox potential, water table depth and soil moisture (using TDR) was used to characterize the site. Significant spatial and temporal variability has been identified, with results from water-table monitoring and some initial chemical analysis from Glastonbury presented here. It appears that during dry periods parts of this site are at risk from desiccation. Analysis of the chemical data, in addition to integrating the results from the other parameters, is ongoing, with the aim of clarifying the risk to the entire site.

Integration of Space and In Situ Observations to Study Global Climate Change

The currently available model-based global data sets of atmospheric circulation are a by-product of the daily requirement of producing initial conditions for numerical weather prediction (NWP) models. These data sets have been quite useful for studying fundamental dynamical and physical processes, and for describing the nature of the general circulation of the atmosphere. However, due to limitations in the early data assimilation systems and inconsistencies caused by numerous model changes, the available model-based global data sets may not be suitable for studying global climate change. A comprehensive analysis of global observations based on a four-dimensional data assimilation system with a realistic physical model should be undertaken to integrate space and in situ observations to produce internally consistent, homogeneous, multivariate data sets for the earth's climate system. The concept is equally applicable for producing data sets for the atmosphere, the oceans, and the biosphere, and such data sets will be quite useful for studying global climate change.

The accuracy of SST retrievals from AATSR: An initial assessment through geophysical validation against in situ radiometers, buoys and other SST data sets

The Advanced Along-Track Scanning Radiometer (AATSR) was launched on Envisat in March 2002. The AATSR instrument is designed to retrieve precise and accurate global sea surface temperature (SST) that, combined with the large data set collected from its predecessors, ATSR and ATSR-2, will provide a long term record of SST data that is greater than 15 years. This record can be used for independent monitoring and detection of climate change. The AATSR validation programme has successfully completed its initial phase. The programme involves validation of the AATSR derived SST values using in situ radiometers, in situ buoys and global SST fields from other data sets. The results of the initial programme presented here will demonstrate that the AATSR instrument is currently close to meeting its scientific objectives of determining global SST to an accuracy of 0.3 K (one sigma). For night time data, the analysis gives a warm bias of between +0.04 K (0.28 K) for buoys to +0.06 K (0.20 K) for radiometers, with slightly higher errors observed for day time data, showing warm biases of between +0.02 (0.39 K) for buoys to +0.11 K (0.33 K) for radiometers. They show that the ATSR series of instruments continues to be the world leader in delivering accurate space-based observations of SST, which is a key climate parameter.

Assessing the diurnal evolution of surface radiation balance over the western region of Tropical Atlantic Ocean using in situ measurements carried out during the FluTuA balance Project

In this work, the diurnal evolution of the radiation balance components over the tropical Atlantic Ocean is described and analysed. The analysis is based on measurements carried Out on board a Brazilian Navy ship during the observational campaign of the FluTuA Project (`Fluxos Turbulentos sobre o Atlantico`), from 15 to 23 May 2002. The observations indicated that the albedo responds its expected to atmospheric attenuation effects with a diurnal evolution similar to the Fresnel albedo. In general, the observed longwave radiation values agreed better with the estimated values obtained without longwave reflection. In agreement with the literature, the average surface emissivity was around 0.97. The net radiation, estimated from published equations for albedo, atmospheric transmissivity and surface emissivity, agreed with the observations, indicating that these parameters are representative of the radiometric properties of the air-sea interface in the region between Natal (6 degrees S, 35.2 degrees W) and the Sao Pedro and Sao Paulo Archipelago (1 degrees N, 29.3 degrees W). Copyright (C) 2008 Royal Meteorological Society

In situ X-ray diffraction studies of phase transition in Pb1-x La x Zr0.40Ti0.60O3 ferroelectric ceramics

The temperature dependence of the crystalline structure and the lattice parameters of Pb1-xLaxZr0.40Ti0.60O3 ferroelectric ceramic system with 0.00 x 0.21 was determined. The samples with x 0.11 show a cubic-to-tetragonal phase transition at the maximum dielectric permittivity, Tmax. Above this amount and especially for the x = 0.12 sample, a spontaneous phase transition from a relaxor ferroelectric state (cubic phase) to a ferroelectric state (tetragonal phase) is observed upon cooling below the Tmax. Unlike what has been reported in other studies, the x = 0.13, 0.14, and 0.15 samples, which present a more pronounced relaxor behavior, also presents a spontaneous normal-to-relaxor transition, indicated by a cubic to tetragonal symmetry below the Tmax. The origin of this anomaly has been associated with an increase in the degree of tetragonality, confirmed by the measurements of the X-ray diffraction patterns. The differential thermal analysis (DSC) measurements also confirm the existence of these phase transitions.

Feasibility of using in situ fusion for the determination of Co, Cr and Mn in Portland cement by direct solid sampling graphite furnace atomic absorption spectrometry

In situ fusion on the boat-type graphite platform has been used as a sample pretreatment for the direct determination of Co, Cr and Mn in Portland cement by solid sampling graphite furnace atomic absorption spectrometry (SS-GF AAS). The 3-field Zeeman technique was adopted for background correction to decrease the sensitivity during measurements. This strategy allowed working with up to 200 mu g of sample. The in situ fusion was accomplished using 10 mu L of a flux mixture 4.0% m/v Na(2)CO(3) + 4.0% m/v ZnO + 0.1% m/v Triton (R) X-100 added over the cement sample and heated at 800 degrees C for 20 s. The resulting mould was completely dissolved with 10 mu L of 0.1% m/v HNO(3). Limits of detection were 0.11 mu g g(-1) for Co, 1.1 mu g g(-1) for Cr and 1.9 mu g g(-1) for Mn. The accuracy of the proposed method has been evaluated by the analysis of certified reference materials. The values found presented no statistically significant differences compared to the certified values (Student`s t-test, p<0.05). In general, the relative standard deviation was lower than 12% (n = 5). (C) 2009 Elsevier B.V. All rights reserved.

Effect of Calcium Pre-rinse and Fluoride Dentifrice on Enamel and on Dental Plaque Formed In Situ

Purpose: The aim of this in situ double-blind randomised crossover study was to investigate the effect of calcium (Ca) pre-rinse on the composition of plaque and on enamel prior to the use of fluoride (F) dentifrice. Materials and Methods: During four phases (14 days each) of this study, 10 volunteers had agreed to wear dental appliances containing two healthy bovine enamel blocks. A fresh solution containing 20% weight/volume (w/v) sucrose was dripped on the enamel blocks ex vivo for 5 min three times a day. Subsequently, the appliances were replaced in the mouth, and the volunteers rinsed their mouth with 10 mL of a Ca (150 mmol/L) or a placebo rinse (1 min). In sequence, a slurry (1:3 w/v) of F (1030 ppm) or placebo dentifrice was dripped onto the blocks ex vivo for 1 min. During this time, the volunteers brushed their teeth with the respective dentifrice. The appliances were replaced in the mouth, and the volunteers rinsed their mouth with water. The plaque formed on the blocks was analysed for F and Ca. The enamel demineralisation as well as the incorporation of F on enamel was evaluated by cross-sectional microhardness and alkali-soluble F analysis, respectively. Data were tested using analysis of variance (P < 0.05). Results: The Ca pre-rinse prior to the use of the F dentifrice led to a three- and sixfold increase in the plaque F and Ca concentrations, respectively. It also did not have any additive effect on the F content on the enamel and the demineralisation of the enamel, in comparison with the use of F dentifrice alone. Conclusions: A Ca lactate rinse used prior to the F dentifrice was able to change the mineral content in the plaque, but it was unable to prevent enamel demineralisation.

Estudo do processo de combustão in-situ em reservatórios maduros de óleos médios e leves (high pressure air injection)

Nearly 3 x 1011 m3 of medium and light oils will remain in reservoirs worldwide after conventional recovery methods have been exhausted and much of this volume would be recovered by Enhanced Oil Recovery (EOR) methods. The in-situ combustion (ISC) is an EOR method in which an oxygen-containing gas is injected into a reservoir where it reacts with the crude oil to create a high-temperature combustion front that is propagated through the reservoir. The High Pressure Air Injection (HPAI) method is a particular denomination of the air injection process applied in light oil reservoirs, for which the combustion reactions are dominant between 150 and 300°C and the generation of flue gas is the main factor to the oil displacement. A simulation model of a homogeneous reservoir was built to study, which was initially undergone to primary production, for 3 years, next by a waterflooding process for 21 more years. At this point, with the mature condition established into the reservoir, three variations of this model were selected, according to the recovery factors (RF) reached, for study the in-situ combustion (HPAI) technique. Next to this, a sensitivity analysis on the RF of characteristic operational parameters of the method was carried out: air injection rate per well, oxygen concentration into the injected gas, patterns of air injection and wells perforations configuration. This analysis, for 10 more years of production time, was performed with assistance of the central composite design. The reservoir behavior and the impacts of chemical reactions parameters and of reservoir particularities on the RF were also evaluated. An economic analysis and a study to maximize the RF of the process were also carried out. The simulation runs were performed in the simulator of thermal processes in reservoirs STARS (Steam, Thermal, and Advanced Processes Reservoir Simulator) from CMG (Computer Modelling Group). The results showed the incremental RF were small and the net present value (NPV) is affected by high initial investments to compress the air. It was noticed that the adoption of high oxygen concentration into the injected gas and of the five spot pattern tends to improve the RF, and the wells perforations configuration has more influence with the increase of the oil thickness. Simulated cases relating to the reservoir particularities showed that smaller residual oil saturations to gas lead to greater RF and the presence of heterogeneities results in important variations on the RF and on the production curves

Estudo dos parâmetros operacionais do processo de combustão in situ em reservatório de petróleo pesado

The occurrence of heavy oil reservoirs have increased substantially and, due to the high viscosity characteristic of this type of oil, conventional recovery methods can not be applied. Thermal methods have been studied for the recovery of this type of oil, with a main objective to reduce its viscosity, by increasing the reservoir temperature, favoring the mobility of the oil and allowing an increasing in the productivity rate of the fields. In situ combustion (ISC) is a thermal recovery method in which heat is produced inside the reservoir by the combustion of part of the oil with injected oxygen, contrasting with the injection of fluid that is heated in the surface for subsequent injection, which leads to loss heat during the trajectory to the reservoir. The ISC is a favorable method for recovery of heavy oil, but it is still difficult to be field implemented. This work had as an objective the parametric analysis of ISC process applied to a semi-synthetic reservoir with characteristics of the Brazilian Northeast reservoirs using vertical production and vertical injection wells, as the air flow injection and the wells completions. For the analysis, was used a commercial program for simulation of oil reservoirs using thermal processes, called Steam, Thermal and Advanced Processes Reservoir Simulator (STARS) from Computer Modelling Group (CMG). From the results it was possible to analyze the efficiency of the ISC process in heavy oil reservoirs by increasing the reservoir temperature, providing a large decrease in oil viscosity, increasing its mobility inside the reservoir, as well as the improvement in the quality of this oil and therefore increasing significantly its recovered fraction. Among the analyzed parameters, the flow rate of air injection was the one which had greater influence in ISC, obtaining higher recovery factor the higher is the flow rate of injection, due to the greater amount of oxygen while ensuring the maintenance of the combustion front

Extração enzimática de óleo e produção in situ de biodiesel a partir da Moringa oleífera Lam

With the growth and development of modern society, arises the need to search for new raw materials and new technologies which present the "clean" characteristic, and do not harm the environment, but can join the energy needs of industry and transportation. The Moringa oleifera Lam, plant originating from India, and currently present in the Brazilian Northeast, presents itself as a multi-purpose plant, can be used as a coagulant in water treatment, as a natural remedy and as a feedstock for biodiesel production. In this work, Moringa has been used as a raw material for studies on the extraction and subsequently in the synthesis of biodiesel. Studies have been conducted on various techniques of Moringa oil extraction (solvents, mechanical pressing and enzymatic), being specially developed an experimental design for the aqueous extraction with the aid of the enzyme Neutrase© 0.8 L, with the aim of analyzing the influence variable pH (5.5-7.5), temperature (45-55°C), time (16-24 hours) and amount of catalyst (2-5%) on the extraction yield. In relation to study of the synthesis of biodiesel was initially carried out a conventional transesterification (50°C, KOH as a catalyst, methanol and 60 minutes reaction). Next, a study was conducted using the technique of in situ transesterification by using an experimental design variables as temperature (30-60°C), catalyst amount (2-5%), and molar ratio oil / ethanol (1:420-1:600). The extraction technique that achieved the highest extraction yield (35%) was the one that used hexane as a solvent. The extraction using 32% ethanol obtained by mechanical pressing and extraction reached 25% yield. For the enzymatic extraction, the experimental design indicated that the extraction yield was most affected by the effect of the combination of temperature and time. The maximum yield obtained in this extraction was 16%. After the step of obtaining the oil was accomplished the synthesis of biodiesel by the conventional method and the in situ technique. The method of conventional transesterification was obtained a content of 100% and esters by in situ technique was also obtained in 100% in the experimental point 7, with a molar ratio oil / alcohol 1:420, Temperature 60°C in 5% weight KOH with the reaction time of 1.5 h. By the experimental design, it was found that the variable that most influenced the ester content was late the percentage of catalyst. By physico-chemical analysis it was observed that the biodiesel produced by the in situ method fell within the rules of the ANP, therefore this technique feasible, because does not require the preliminary stage of oil extraction and achieves high levels of esters

Avaliação da produção de biodiesel de microalga Isochrysis galbana via transesterificação in situ

Microalgae are microscopic photosynthetic organisms that grow rapidly and in different environmental conditions due to their simple cellular structure. The cultivation of microalgae is a biological system capable of storing solar energy through the production of organic compounds via photosynthesis, and these species presents growth faster than land plants, enabling higher biomass yield. Thus, it is understood that the cultivation of these photosynthetic mechanisms is part of a relevant proposal, since, when compared to other oil producing raw materials, they have a significantly higher productivity, thus being a raw material able to complete the current demand by biodiesel . The overall aim of the thesis was to obtain biofuel via transesterification process of bio oil from the microalgae Isochrysis galbana. The specific objective was to estimate the use of a photobioreactor at the laboratory level, for the experiments of microalgae growth; evaluating the characteristics of biodiesel from microalgae produced by in situ transesterification process; studying a new route for disinfection of microalgae cultivation, through the use of the chemical agent sodium hypochlorite. The introduction of this new method allowed obtaining the kinetics of the photobioreactor for cultivation, besides getting the biomass needed for processing and analysis of experiments in obtaining biodiesel. The research showed acceptable results for the characteristics observed in the bio oil obtained, which fell within the standards of ANP Resolution No. 14, dated 11.5.2012 - 18.5.2012. Furthermore, it was demonstrated that the photobioreactor designed meet expectations about study culture growth and has contributed largely to the development of the chosen species of microalgae. Thus, it can be seen that the microalgae Isochrysis galbana showed a species with potential for biodiesel production

Caracterização in situ e diversidade genética de algodoeiros mocós (Gossypium hissutum raça marie galante) da Região Nordeste do Brasil

Brazil is one of the major centers of diversity for polyploid cotton plants; these plants belong to the genus Gossypium, which has three known species: G. hirsutum, G. barbadense and G. mustelinum. The Northeast is the only region where the three species occur, the last group being endemic. Northeast s cotton plants can be important sources of variability for genetic breeding. It is believed that great part of local diversity is being lost, due to economic, political, cultural and agricultural problems. In an attempt to mitigate this loss and delineate conservation strategies it is necessary to know how the species are found where they occur. The objective was to characterize and determine how plants are maintained in situ in the states of Maranhão, Piauí, Ceará, Rio Grande do Norte and Paraíba at the beginning of the XXI century. The in situ characterization of G. hirsutum and G. barbadense was conducted through structured interviews with the cotton plants owners and through the analysis of the environment. The data were collected during expeditions undertaken between the years 2004 to 2005. Twenty-two plants were collected in the state of Paraíba, forty-four in the state of Rio Grande do Norte, one hundred and forty-six in the state of Ceará, forty in the state of Maranhão and ninety-one plants in the state of Piauí. All plants collected in the states of Paraíba and Rio Grande do Norte belonged to moco type. Moco cotton plants also predominated in the other states, representing 92%, 62% and 78% of plants collected in Ceará, Piauí and Maranhão, respectively. The other cotton plants collected belong to the species G. barbadense. The cotton plants were found in situ as dooryard plants, roads side, feral populations, cultivation or local varieties. Great part were dooryard plants (45.2%), being major in Piauí and Maranhão. Cultivation predominated in Ceará; in Rio Grande do Norte feral populations were the most frequent and, in Paraíba, local varieties. The maintenance of moco plants is related, mainly, to the phytotherapic domestic use (20.9%) and to confection of lamp wicks (29.7%). Few inhabitants in Paraíba, Rio Grande do Norte, Piauí and none in Maranhão used harvest the plants, storage the seeds or gin; however, in Ceará, 40.5% of owners affirmed that they harvested and commercialized the fiber. It was found that the maintenance of species is dependent of the fragile cultural habits of local inhabitants, therefore the maintenance in situ is not a suitable way to conservation of genetic resources. The efforts must be directed to the continuity of collections, maintenance and characterization ex situ

Borderline leprosy: in situ and cytokine profile in supernatant of mononuclear of cell culture

In order to contribute to a better understanding of cytokine participation in borderline leprosy, in the present study we determined - by in vitro and in situ examinations - the production of these cytokine mediation in non-treated borderline tuberculoid (BT) patients and borderline lepromatous (BL) patients. Seven non-treated BT patients, 12 non-treated BL patients, besides 19 healthy individuals (control group), were evaluated. Peripheral blood mononuclear cells (PBMC) were stimulated or not with specific-M. leprae stimulus (whole and sonicated M. leprae antigens) and a non-specific stimulus. After 48 hours, supernatant was collected for TNF-alpha, IFN-gamma, IL-10 and TGF-beta1 cytokine determination by ELISA. Biopsies from cutaneous lesions were submitted to histological analysis and hematoxylin-eosin and Fite-Faraco stainings; the sections then underwent iNOS, IL-10 and TGF-beta1 in situ detection by immunohistochemistry. Cytokine quantification in PBMC supernatants from patients showed that BT patients produced higher levels of IFN-gamma. Compared to healthy individuals, both borderline patient groups produced lower levels of TGF-beta1 while BL patients generated lower IL-10 levels. The in situ iNOS expression was higher in BT patients compared to BL individuals. on the order hand, TGF-beta1 cytokine revealed a higher proportion of immunostained cells in BL patients. There was no significant difference in IL-10 level between BT and BL patients. Regarding cutaneous lesions, in BL patients there was a negative correlation between TGF-beta1 tissue expression and IL-10. Independently of the clinical form, we observed a positive correlation between TGF-beta1 and bacterial index as well as a negative correlation between the TGF-beta1 tissue expression and iNOS. The results even showed a positive correlation between iNOS tissue expression and production of IFN-gamma by PBMC stimulated with M. leprae antigens. Taken together, the histopathological and immunological observations reinforce the notion of immunological instability in borderline leprosy patients and indicating the participation of mixed cytokines profiles in these individuals, specifically a Th1 profile in BT patients and Th2 profile in BL patients, with a possible participation of T-regulatory lymphocytes.

A novel in situ Polymerase chain reaction hybridisation assay for the direct detection of bovine herpesvirus type 5 in formalin-fixed, paraffin-embedded tissues

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)