Molecular screening of 246 Portuguese Aspergillus isolates among different clinical and environmental sources

Clinical and environmental samples from Portugal were screened for the presence of Aspergillus and the distributions of the species complexes were determined in order to understand how their distributions differ based on their source. Fifty-seven Aspergillus isolates from clinical samples were collected from 10 health institutions. Six species complexes were detected by internal transcribed spacer sequencing; Fumigati, Flavi, and Nigri were found most frequently (50.9%, 21.0%, and 15.8%, respectively). β-tubulin and calmodulin sequencing resulted in seven cryptic species (A. awamorii, A. brasiliensis, A. fructus, A. lentulus, A. sydowii, A. tubingensis, Emericella echinulata) being identified among the 57 isolates. Thirty-nine isolates of Aspergillus were recovered from beach sand and poultry farms, 31 from swine farms, and 80 from hospital environments, for a total 189 isolates. Eleven species complexes were found in these 189 isolates, and those belonging to the Versicolores species complex were found most frequently (23.8%). There was a significant association between the different environmental sources and distribution of the species complexes; the hospital environment had greater variability of species complexes than other environmental locations. A high prevalence of cryptic species within the Circumdati complex was detected in several environments; from the isolates analyzed, at least four cryptic species were identified, most of them growing at 37ºC. Because Aspergillus species complexes have different susceptibilities to antifungals, knowing the species-complex epidemiology for each setting, as well as the identification of cryptic species among the collected clinical isolates, is important. This may allow preventive and corrective measures to be taken, which may result in decreased exposure to those organisms and a better prognosis.

Biaxial Nematic Order in the Hard-boomerang Fluid

We consider a fluid of hard boomerangs, each composed of two hard spherocylinders joined at their ends at an angle Psi. The resulting particle is nonconvex and biaxial. The occurence of nematic order in such a system has been investigated using Straley's theory, which is a simplificaton of Onsager's second-virial treatment of long hard rods, and by bifurcation analysis. The excluded volume of two hard boomerangs has been approximated by the sum of excluded volumes of pairs of constituent spherocylinders, and the angle-dependent second-virial coefficient has been replaced by a low-order interpolating function. At the so-called Landau point, Psi(Landau)approximate to 107.4 degrees, the fluid undergoes a continuous transition from the isotropic to a biaxial nematic (B) phase. For Psi not equal Psi(Landau) ordering is via a first-order transition into a rod-like uniaxial nematic phase (N(+)) if Psi > Psi(Landau), or a plate-like uniaxial nematic (N(-)) phase if Psi < Psi(Landau). The B phase is separated from the N(+) and N(-) phases by two lines of continuous transitions meeting at the Landau point. This topology of the phase diagram is in agreement with previous studies of spheroplatelets and biaxial ellipsoids. We have checked the accuracy of our theory by performing numerical calculations of the angle-dependent second virial coefficient, which yields Psi(Landau)approximate to 110 degrees for very long rods, and Psi(Landau)approximate to 90 degrees for short rods. In the latter case, the I-N transitions occur at unphysically high packing fractions, reflecting the inappropriateness of the second-virial approximation in this limit.

Comparação da fixação em formalina e em fixador molecular e sua influência em técnicas histoquímicas em tecido hepático

A Formalina 10% Tamponada (FNT 10%) é considerada o fixador de eleição nos Laboratórios de Anatomia Patológica. Contudo, a exposição a esta substância acarreta riscos para a saúde dos técnicos. A International Agency for Research on Cancer (IARC) classificou-a como cancerígeno humano e estudos relevam uma correlação positiva entre a exposição a formaldeído e o desenvolvimento de leucemia e leucemia mielóide. Torna-se relevante alterar o processo de fixação substituindo a FNT 10% por outros fixadores melhorando estas questões. Como tal, desenvolveram-se fixadores à base de outros compostos químicos que não formaldeído, como o Fixador Molecular Universal (UMFIX) que tem como base metanol e polietilenoglicol e que é usualmente utilizado com um processador rápido de micro-ondas. Pretende-se comparar as diferenças entre a fixação por FNT 10% e a fixação por UMFIX, em tecido hepático processado em processador rápido de micro-ondas, para as colorações de Hematoxilina-Eosina (H&E), Tricrómio de Gomori, Reticulina e PAS, através da qualidade final das lâminas testadas. A coloração de H&E foi a única que apresentou diferenças estatisticamente significativas entre os dois fixadores (p=0,032; α<0,05; Mann-Whitney). O UMFIX apresenta-se como um substituto da FNT 10% para o processamento rápido em micro-ondas, pois além de apresentar lâminas com uma qualidade final semelhante ou superior às da FNT 10%, ultrapassa os riscos referidos.

Fast growing fungi: a problem to be solved to achieve characterization of occupational exposure to fungi in cork industry

Chrysonilia sitophila is a common mould in cork industry and has been identified as a cause of IgE sensitization and occupational asthma. This fungal species have a fast growth rate that may inhibit others species’ growth causing underestimated data from characterization of occupational fungal exposure. Aiming to ascertain occupational exposure to fungi in cork industry, were analyzed papers from 2000 about the best air sampling method, to obtain quantification and identification of all airborne culturable fungi, besides the ones that have fast-growing rates. Impaction method don’t allows the collection of a representative air volume, because even with some media that restricts the growth of the colonies, in environments with higher fungal load, such as cork industry, the counting of the colonies is very difficult. Otherwise, impinger method permits the collection of a representative air volume, since we can make dilution of the collected volume. Besides culture methods that allows fungal identification trough macro- and micro-morphology, growth features, thermotolerance and ecological data, we can apply molecular biology with the impinger method, to detect the presence of non-viable particles and potential mycotoxin producers’ strains, and also to detect mycotoxins presence with ELISA or HPLC. Selection of the best air sampling method in each setting is crucial to achieve characterization of occupational exposure to fungi. Information about the prevalent fungal species in each setting and also the eventual fungal load it’s needed for a criterious selection.

Molecular epidemiology of Aspergillus collected from cystic fibrosis patients

Background - Aspergillus respiratory infection is a common complication in cystic fibrosis (CF) and is associated with loss of pulmonary function and allergic disease. Methods - Fifty-three Aspergillus isolates recovered from CF patients were identified to species by Internal Transcribed Spacer Region (ITS), β-tubulin, and calmodulin sequencing. Results - Three species complexes (Terrei, Nigri, and Fumigati) were found. Identification to species level gave a single Aspergillus terreus sensu stricto, one Aspergillus niger sensu stricto and 51 Aspergillus fumigatus sensu stricto isolates. No cryptic species were found. Conclusions - To our knowledge, this is the first prospective study of Aspergillus species in CF using molecular methods. The paucity of non-A. fumigatus and of cryptic species of A. fumigatus suggests a special association of A. fumigatus sensu stricto with CF airways, indicating it likely displays unique characteristics making it suitable for chronic residence in that milieu. These findings could refine an epidemiologic and therapeutic approach geared to this pathogen.

Avaliação da distribuição de dose em doentes com cancro da mama: cobertura e homogeneidade no volume alvo e dose nos órgãos de risco

Mestrado em Radioterapia

Aurícula direita: mecânica e sobrecarga de volume

Mestrado em Tecnologia de Diagnóstico e Intervenção Cardiovascular - Ramo de especialização: Ultrassonografia Cardiovascular

Potential pathogenic fungi assessment through molecular biology in cork industry

Portugal has been the world leader in the cork sector in terms of exports, employing ten thousands of workers. In this working activity, the permanent contact with cork may lead to the exposure to fungi, raising concerns as potential occupational hazards in cork industry. The application of molecular tools is crucial in this setting, since fungal species with faster growth rates may hide other species with clinical relevance, such as species belonging to P. glabrum and A. fumigatus complexes. A study was developed aiming at assessing fungal contamination due to Aspergillus fumigatus complex and Penicillium glabrum complex by molecular methods in three cork industries in the outskirt of Lisbon city.

Molecular mobility, composition and structure analysis in glycerol plasticised chitosan films

This study was developed with the purpose to investigate the effect of polysaccharide/plasticiser concentration on the microstructure and molecular dynamics of polymeric film systems, using transmission electron microscope imaging (TEM) and nuclear magnetic resonance (NMR) techniques. Experiments were carried out in chitosan/glycerol films prepared with solutions of different composition. The films obtained after drying and equilibration were characterised in terms of composition, thickness and water activity. Results show that glycerol quantities used in film forming solutions were responsible for films composition; while polymer/total plasticiser ratio in the solution determined the thickness (and thus structure) of the films. These results were confirmed by TEM. NMR allowed understanding the films molecular rearrangement. Two different behaviours for the two components analysed, water and glycerol were observed: the first is predominantly moving free in the matrix, while glycerol is mainly bounded to the chitosan chain. (C) 2013 Elsevier Ltd. All rights reserved.

Densities and refractive indices for the ternary mixture methanol/propan-1-Ol/acetonitrile

Refractive indices, n(D), and densities, rho, at 298.15 K were measured for the ternary mixture methanol (MeOH)/propan-1-ol (1-PrOH)/acetonitrile (MeCN) for a total of 22 mole fractions, along with 18 mole fractions of each of the corresponding binary mixtures, methanol/propan-1-ol, propan-1-ol/acetonitrile and methanol/acetonitrile. The variation of excess refractive indices and excess molar volumes with composition was modeled by the Redlich-Kister polynomial function in the case of binary mixtures and by the Cibulka equation for the ternary mixture. A thermodynamic approach to excess refractive indices, recently proposed by other authors, was applied for the first time to ternary liquid mixtures. Structural effects were identified and interpreted both in the binary and ternary systems. A complex relationship between excess refractive indices and excess molar volumes was identified, revealing all four possible sign combinations between these two properties. Structuring of the mixtures was also discussed on the basis of partial molar volumes of the binary and ternary mixtures.

A cellulosic liquid crystal pool for cellulose nanocrystals: structure and molecular dynamics at high shear rates

Cellulose and its derivatives, such as hydroxypropylcellulose (HPC) have been studied for a long time but they are still not well understood particularly in liquid crystalline solutions. These systems can be at the origin of networks with properties similar to liquid crystalline (LC) elastomers. The films produced from LC solutions can be manipulated by the action of moisture allowing for instance the development of a soft motor (Geng et al., 2013) driven by humidity. Cellulose nanocrystals (CNC), which combine cellulose properties with the specific characteristics of nanoscale materials, have been mainly studied for their potential as a reinforcing agent. Suspensions of CNC can also self-order originating a liquid-crystalline chiral nematic phases. Considering the liquid crystalline features that both LC-HPC and CNC can acquire, we prepared LC-HPC/CNC solutions with different CNC contents (1,2 and 5 wt.%). The effect of the CNC into the LC-HPC matrix was determined by coupling rheology and NMR spectroscopy - Rheo-NMR a technique tailored to analyse orientational order in sheared systems. (C) 2015 Elsevier Ltd. All rights reserved.

Molecular fingerprint of human gastric cell line infected by Helicobacter pylori

Helicobacter pylori infection represents a serious health problem, given its association with serious gastric diseases as gastric ulcers, cancer and MALT lymphoma. Currently no vaccine exists and antibiotic-based eradication therapy is already failing in more than 20% of cases. To increase the knowledge on the infection process diverse gastric cell lines, e.g. the adenocarcinona gastric (AGS) cell line, are routinely used has in vitro models of gastric epithelia. In the present work the molecular fingerprint of infected and non-infected AGS cell lines, by diverse H. pylori strains, was acquired using vibrational infrared spectroscopy. These molecular fingerprints enabled to discriminate infected from non-infected AGS cells, and infection due to different strains, by performing Principal Component Analysis. It was also possible to estimate, from the AGS cells molecular fingerprint, the effect of the infection on diverse biochemical and metabolic cellular status. In resume infra-red spectroscopy enabled the acquisition of infected AGS cells molecular fingerprint with minimal sample preparation, in a rapid, high-throughput, economic process yielding highly sensitive and informative data, most useful for promoting critical knowledge on the H. pylori infection process. © 2015 IEEE.

Molecular details of INH-C-10 binding to wt KatG and Its S315T mutant

Isoniazid (INH) is still one of the two most effective antitubercular drugs and is included in all recommended multitherapeutic regimens. Because of the increasing resistance of Mycobacterium tuberculosis to INH, mainly associated with mutations in the katG gene, new INH-based compounds have been proposed to circumvent this problem. In this work, we present a detailed comparative study of the molecular determinants of the interactions between wt KatG or its S315T mutant form and either INH or INH-C10, a new acylated INH derivative. MD simulations were used to explore the conformational space of both proteins, and results indicate that the S315T mutation did not have a significant impact on the average size of the access tunnel in the vicinity of these residues. Our simulations also indicate that the steric hindrance role assigned to Asp137 is transient and that electrostatic changes can be important in understanding the enzyme activity data of mutations in KatG. Additionally, molecular docking studies were used to determine the preferred modes of binding of the two substrates. Upon mutation, the apparently less favored docking solution for reaction became the most abundant, suggesting that S315T mutation favors less optimal binding modes. Moreover, the aliphatic tail in INH-C10 seems to bring the hydrazine group closer to the heme, thus favoring the apparent most reactive binding mode, regardless of the enzyme form. The ITC data is in agreement with our interpretation of the C10 alkyl chain role and helped to rationalize the significantly lower experimental MIC value observed for INH-C10. This compound seems to be able to counterbalance most of the conformational restrictions introduced by the mutation, which are thought to be responsible for the decrease in INH activity in the mutated strain. Therefore, INH-C10 appears to be a very promising lead compound for drug development.

Viscosity and self-diffusion coefficients of dialkyl adipates: a correlation scheme with predictive capabilities

A correlation and predictive scheme for the viscosity and self-diffusivity of liquid dialkyl adipates is presented. The scheme is based on the kinetic theory for dense hard-sphere fluids, applied to the van der Waals model of a liquid to predict the transport properties. A "universal" curve for a dimensionless viscosity of dialkyl adipates was obtained using recently published experimental viscosity and density data of compressed liquid dimethyl (DMA), dipropyl (DPA), and dibutyl (DBA) adipates. The experimental data are described by the correlation scheme with a root-mean-square deviation of +/- 0.34 %. The parameters describing the temperature dependence of the characteristic volume, V-0, and the roughness parameter, R-eta, for each adipate are well correlated with one single molecular parameter. Recently published experimental self-diffusion coefficients of the same set of liquid dialkyl adipates at atmospheric pressure were correlated using the characteristic volumes obtained from the viscosity data. The roughness factors, R-D, are well correlated with the same single molecular parameter found for viscosity. The root-mean-square deviation of the data from the correlation is less than 1.07 %. Tests are presented in order to assess the capability of the correlation scheme to estimate the viscosity of compressed liquid diethyl adipate (DEA) in a range of temperatures and pressures by comparison with literature data and of its self-diffusivity at atmospheric pressure in a range of temperatures. It is noteworthy that no data for DEA were used to build the correlation scheme. The deviations encountered between predicted and experimental data for the viscosity and self-diffusivity do not exceed 2.0 % and 2.2 %, respectively, which are commensurate with the estimated experimental measurement uncertainty, in both cases.