Direct visualization of Ras proteins in spatially distinct cell surface microdomains

Localization of signaling complexes to specific micro-domains coordinates signal transduction at the plasma membrane. Using immunogold electron microscopy of plasma membrane sheets coupled with spatial point pattern analysis, we have visualized morphologically featureless microdomains including lipid rafts, in situ and at high resolution. We find that an inner-plasma membrane lipid raft marker displays cholesterol-dependent clustering in microdomains with a mean diameter of 44 nm that occupy 35% of the cell surface. Cross-linking an outer-leaflet raft protein results in the redistribution of inner leaflet rafts, but they retain their modular structure. Analysis of Ras microlocalization shows that inactive H-ras is distributed between lipid rafts and a cholesterol-independent micro-domain. Conversely, activated H-ras and K-ras reside predominantly in nonoverlapping, cholesterol-independent microdomains. Galectin-1 stabilizes the association of activated H-ras with these nonraft microdomains, whereas K-ras clustering is supported by farnesylation, but not geranylgeranylation. These results illustrate that the inner plasma membrane comprises a complex mosaic of discrete microdomains. Differential spatial localization within this framework can likely account for the distinct signal outputs from the highly homologous Ras proteins.

A comparison of two methods for the calculation of accumulated oxygen deficit

The aim of this study was to compare accumulated oxygen deficit data derived using two different exercise protocols with the aim of producing a less time-consuming test specifically for use with athletes. Six road and four track male endurance cyclists performed two series of cycle ergometer tests. The first series involved five 10 min sub-maximal cycle exercise bouts, a (V) over dotO(2peak) test and a 115% (V) over dotO(2peak) test. Data from these tests were used to estimate the accumulated oxygen deficit according to the calculations of Medbo et al. (1988). In the second series of tests, participants performed a 15 min incremental cycle ergometer test followed, 2 min later, by a 2 min variable resistance test in which they completed as much work as possible while pedalling at a constant rate. Analysis revealed that the accumulated oxygen deficit calculated from the first series of tests was higher (P< 0.02) than that calculated from the second series: 52.3 +/- 11.7 and 43.9 +/- 6.4 ml . kg(-1), respectively (mean +/- s). Other significant differences between the two protocols were observed for (V) over dot O-2peak, total work and maximal heart rate; all were higher during the modified protocol (P

Comparison of different methods for the determination of total organic carbon and humic substances in Brazilian soils

ABSTRACTAiming to compare three different methods for the determination of organic carbon (OC) in the soil and fractions of humic substances, seventeen Brazilian soil samples of different classes and textures were evaluated. Amounts of OC in the soil samples and the humic fractions were measured by the dichromate-oxidation method, with and without external heating in a digestion block at 130 °C for 30 min; by the loss-on-ignition method at 450 °C during 5 h and at 600 °C during 6 h; and by the dry combustion method. Dry combustion was used as reference in order to measure the efficiency of the other methods. Soil OC measured by the dichromate-oxidation method with external heating had the highest efficiency and the best results comparing to the reference method. When external heating was not used, the mean recovery efficiency dropped to 71%. The amount of OC was overestimated by the loss-on-ignition methods. Regression equations obtained between total OC contents of the reference method and those of the other methods showed relatively good adjustment, but all intercepts were different from zero (p < 0.01), which suggests that more accuracy can be obtained using not one single correction factor, but considering also the intercept. The Walkley-Black method underestimated the OC contents of the humic fractions, which was associated with the partial oxidation of the humin fraction. Better results were obtained when external heating was used. For the organic matter fractions, the OC in the humic and fulvic acid fractions can be determined without external heating if the reference method is not available, but the humin fraction requires the external heating.

Analysis and visualization of chromosome information

This paper analyzes the DNA code of several species in the perspective of information content. For that purpose several concepts and mathematical tools are selected towards establishing a quantitative method without a priori distorting the alphabet represented by the sequence of DNA bases. The synergies of associating Gray code, histogram characterization and multidimensional scaling visualization lead to a collection of plots with a categorical representation of species and chromosomes.

Mould and yeast identification in archival settings: preliminary results on the use of traditional methods and molecular biology options in Portuguese archives

This project was developed to fully assess the indoor air quality in archives and libraries from a fungal flora point of view. It uses classical methodologies such as traditional culture media – for the viable fungi – and modern molecular biology protocols, especially relevant to assess the non-viable fraction of the biological contaminants. Denaturing high-performance liquid chromatography (DHPLC) has emerged as an alternative to denaturing gradient gel electrophoresis (DGGE) and has already been applied to the study of a few bacterial communities. We propose the application of DHPLC to the study of fungal colonization on paper-based archive materials. This technology allows for the identification of each component of a mixture of fungi based on their genetic variation. In a highly complex mixture of microbial DNA this method can be used simply to study the population dynamics, and it also allows for sample fraction collection, which can, in many cases, be immediately sequenced, circumventing the need for cloning. Some examples of the methodological application are shown. Also applied is fragment length analysis for the study of mixed Candida samples. Both of these methods can later be applied in various fields, such as clinical and sand sample analysis. So far, the environmental analyses have been extremely useful to determine potentially pathogenic/toxinogenic fungi such as Stachybotrys sp., Aspergillus niger, Aspergillus fumigatus, and Fusarium sp. This work will hopefully lead to more accurate evaluation of environmental conditions for both human health and the preservation of documents.

Efficiency of sampling methods and effort to assess arthropod diversity in Azorean native forests

The BALA project (Biodiversity of Arthropods of Laurisilva of the Azores) is a research initiative to quantify the spatial distribution of arthropod biodiversity in native forests of the Azores archipelago. Arthropods were collected using a combination of two techniques, targeting epigean (ground dwelling) and canopy (arboreal) arthropods: pitfall traps (with Turquin and Ethylene solutions) and beating samples (using the three most dominant plant species). A total of 109 transects distributed amongst 18 forest fragments in seven of the nine Azorean islands were used in this study. The performance of alternative sampling methods and effort were tested. No significant differences were found in the accumulated number of species captured whether an alternative method was used or whether another transect with similar effort was established in another location within the same fragment. A combination of Ethylene and Turquin traps captured more species per individual, Turquin and beating captured more species per sample, and Turquin captured more species per unit time. An optimization exercise was performed and we found that the protocol applied during recent years is very close to optimal, allowing its future replication with confidence. The minimum combinations of sampling effort and methods, in order to monitor or to inventory diversity, taking into account different proportions of sample completeness are discussed.

Development of electrochemical methods for determination of tramadol - analytical application to pharmaceutical dosage forms

A square-wave voltammetric (SWV) method and a flow injection analysis system with amperometric detection were developed for the determination of tramadol hydrochloride. The SWV method enables the determination of tramadol over the concentration range of 15-75 µM with a detection limit of 2.2 µM. Tramadol could be determined in concentrations between 9 and 50 µM at a sampling rate of 90 h-1, with a detection limit of 1.7 µM using the flow injection system. The electrochemical methods developed were successfully applied to the determination of tramadol in pharmaceutical dosage forms, without any pre-treatment of the samples. Recovery trials were performed to assess the accuracy of the results; the values were between 97 and 102% for both methods.

Fractional dynamics and MDS visualization of earthquake phenomena

This paper analyses earthquake data in the perspective of dynamical systems and fractional calculus (FC). This new standpoint uses Multidimensional Scaling (MDS) as a powerful clustering and visualization tool. FC extends the concepts of integrals and derivatives to non-integer and complex orders. MDS is a technique that produces spatial or geometric representations of complex objects, such that those objects that are perceived to be similar in some sense are placed on the MDS maps forming clusters. In this study, over three million seismic occurrences, covering the period from January 1, 1904 up to March 14, 2012 are analysed. The events are characterized by their magnitude and spatiotemporal distributions and are divided into fifty groups, according to the Flinn–Engdahl (F–E) seismic regions of Earth. Several correlation indices are proposed to quantify the similarities among regions. MDS maps are proven as an intuitive and useful visual representation of the complex relationships that are present among seismic events, which may not be perceived on traditional geographic maps. Therefore, MDS constitutes a valid alternative to classic visualization tools for understanding the global behaviour of earthquakes.

Visualization of clusters in geo-referenced data using three-dimensional self-organizing maps

Dissertação apresentada como requisito parcial para obtenção do grau de Mestre em Estatística e Gestão de Informação

Applications of Fourier methods to the analysis of some stochastic processes

Dissertação de Doutoramento em Matemática: Processos Estocásticos

Diagnosis of Helicobacter pylori: comparison of an urease test, histological visualization of curved bacteria and culture

Helicobacter pylori was investigated in 189 patients for culture, microscopic visualization of campylobacter-like organisms (CLO) and a ten minute urease test. In 136 (72%) the bacteria was isolated, and in 98 of them CLO were histologically detected. Specificity, sensitivity, positive and negative predictive values of microscopic visualization of CLO were: 0.77, 0.73, 0.97 and 0.51, respectively; 98 culture-positive patients were urease test positive. Specificity, sensitivity, positive and negative predictive values of the urease test were: 0.83, 0.72, 0.92 and 0.54, respectively. Comparing the urease test with culture of H. pylori combined with microscopic visualization of CLO, its specificity, sensitivity, positive and negative predictive values were: 0.95, 0.71, 0.98 and 0.48, respectively. Probably, these values are not real, since bacteria different from H. pylori could be misclassified as CLO.

Multidimensional scaling visualization of earthquake phenomena

Earthquakes are associated with negative events, such as large number of casualties, destruction of buildings and infrastructures, or emergence of tsunamis. In this paper, we apply the Multidimensional Scaling (MDS) analysis to earthquake data. MDS is a set of techniques that produce spatial or geometric representations of complex objects, such that, objects perceived to be similar/distinct in some sense are placed nearby/distant on the MDS maps. The interpretation of the charts is based on the resulting clusters since MDS produces a different locus for each similarity measure. In this study, over three million seismic occurrences, covering the period from January 1, 1904 up to March 14, 2012 are analyzed. The events, characterized by their magnitude and spatiotemporal distributions, are divided into groups, either according to the Flinn–Engdahl seismic regions of Earth or using a rectangular grid based in latitude and longitude coordinates. Space-time and Space-frequency correlation indices are proposed to quantify the similarities among events. MDS has the advantage of avoiding sensitivity to the non-uniform spatial distribution of seismic data, resulting from poorly instrumented areas, and is well suited for accessing dynamics of complex systems. MDS maps are proven as an intuitive and useful visual representation of the complex relationships that are present among seismic events, which may not be perceived on traditional geographic maps. Therefore, MDS constitutes a valid alternative to classic visualization tools, for understanding the global behavior of earthquakes.

Dynamic Analysis and Pattern Visualization of Forest Fires

This paper analyses forest fires in the perspective of dynamical systems. Forest fires exhibit complex correlations in size, space and time, revealing features often present in complex systems, such as the absence of a characteristic length-scale, or the emergence of long range correlations and persistent memory. This study addresses a public domain forest fires catalogue, containing information of events for Portugal, during the period from 1980 up to 2012. The data is analysed in an annual basis, modelling the occurrences as sequences of Dirac impulses with amplitude proportional to the burnt area. First, we consider mutual information to correlate annual patterns. We use visualization trees, generated by hierarchical clustering algorithms, in order to compare and to extract relationships among the data. Second, we adopt the Multidimensional Scaling (MDS) visualization tool. MDS generates maps where each object corresponds to a point. Objects that are perceived to be similar to each other are placed on the map forming clusters. The results are analysed in order to extract relationships among the data and to identify forest fire patterns.