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.

Novel potentiometric sensors of molecular imprinted polymers for specific binding of chlormequat

Molecularly imprinted polymers (MIP) were used as potentiometric sensors for the selective recognition and determination of chlormequat (CMQ). They were produced after radical polymerization of 4-vinyl pyridine (4-VP) or methacrylic acid (MAA) monomers in the presence of a cross-linker. CMQwas used as template. Similar nonimprinted (NI) polymers (NIP) were produced by removing the template from reaction media. The effect of kind and amount of MIP or NIP sensors on the potentiometric behavior was investigated. Main analytical features were evaluated in steady and flow modes of operation. The sensor MIP/4-VP exhibited the best performance, presenting fast near-Nernstian response for CMQover the concentration range 6.2×10-6 – 1.0×10-2 mol L-1 with detection limits of 4.1×10-6 mol L-1. The sensor was independent from the pH of test solutions in the range 5 – 10. Potentiometric selectivity coefficients of the proposed sensors were evaluated over several inorganic and organic cations. Results pointed out a good selectivity to CMQ. The sensor was applied to the potentiometric determination of CMQin commercial phytopharmaceuticals and spiked water samples. Recoveries ranged 96 to 108.5%.

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.

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.

Contributo para a validação de uma escala pediátrica, Peabody Developmental Motor Scale-2 (versão portuguesa)

Mestrado em Fisioterapia

Caracterização molecular da resistência a antimicrobianos em Escherichia coli produtoras de β-lactamases de espectro alargado isoladas em cães e estudo da partilha de clones bacterianos entre animais de companhia e coabitantes humanos

A pressão seletiva originada pelo uso excessivo de antimicrobianos na medicina humana e veterinária tem contribuído para a emergência de estirpes bacterianas multirresistentes, sendo os estudos mais escassos relativamente à sua presença nos animais de companhia. Porque os animais e os seus proprietários partilham o mesmo espaço habitacional, apresentando comportamentos de contacto próximo, existe uma hipótese elevada de transferência microbiana inter-espécie. Ante esta possibilidade é importante escrutinar o papel dos animais de companhia enquanto reservatórios de estirpes e de genes de resistência, bem como a sua envolvência na disseminação de estirpes bacterianas multirresistentes. Importa também, investigar o papel das superfícies e objetos domésticos partilhados por ambos, como potenciadores deste fenómeno. O objetivo deste trabalho foi, identificar o filogrupo e fazer a caracterização molecular dos genes que conferem resistência aos β-lactâmicos e às quinolonas, em quarenta isolados de Escherichia coli produtoras de β-lactamases de espectro alargado (ESBL), obtidas em zaragatoas fecais de cães consultados no Hospital Veterinário do ICBAS-UP. Complementarmente pretendeu-se inferir sobre a partilha de clones de Escherichia coli e Enterococcus spp. com elevadas resistências, em cinco agregados familiares (humanos e seus animais de companhia) assim como avaliar a potencial disseminação de estirpes multirresistentes no ambiente doméstico. Previamente foram recolhidas zaragatoas de fezes, pelo e mucosa oral dos animais e em alguns casos, dos seus proprietários, e ainda do ambiente doméstico. As zaragatoas foram processadas e as estirpes isoladas com base em meios seletivos. Foram realizados testes de suscetibilidade antimicrobiana de modo a estabelecer o fenótipo de resistência de cada isolado. O DNA foi extraído por varias metodologias e técnicas de PCR foram utilizadas para caracterização de filogrupos (Escherichia coli) e identificação da espécie (Enterococcus spp.). A avaliação da proximidade filogenética entre isolados foi efetuada por ERIC PCR e PFGE. No conjunto de quarenta isolados produtores de ESBL e/ou resistentes a quinolonas verificou-se que 47,5% pertenciam ao filogrupo A, havendo uma menor prevalência do filogrupo D (25,0%), B1 (17,5%), e B2 (10,0%).A frequência de resistência nestes isolados é factualmente elevada, sendo reveladora de uma elevada pressão seletiva. Com exceção de dois isolados, os fenótipos foram justificados pela presença de β-lactamases. A frequência da presença de genes foi: 47% blaTEM, 34% blaSHV, 24% blaOXA , 18% blaCTX-M-15, 8% blaCTX-M-2, 3% blaCTX-M-9. Nos isolados resistentes às quinolonas verificou-se maioritariamente a presença de mutações nos genes cromossomais gyrA e parC, e em alguns casos a presença de um determinante de resistência mediado por plasmídeo – qnrS. Nos cinco “agregados familiares” (humanos e animais) estudados foi observada uma partilha frequente de clones de E. coli e Enterococcus faecalis com múltiplas resistências, isolados em fezes e mucosa oral de cães e gatos e fezes e mãos dos respetivos proprietários, evidenciando-se assim uma possível transferência direta entre coabitantes (agregados A, C, D, E). Ficou também comprovado com percentagens de similaridade genotípica superiores a 94% que essa disseminação também ocorre para o ambiente doméstico, envolvendo objetos dos animais e de uso comum (agregados A, E). Os resultados obtidos reforçam a necessidade de um uso prudente dos antimicrobianos, pois elevados padrões de resistências terão um impacto não só na qualidade de vida dos animais mas também na saúde humana. Adicionalmente importa sensibilizar os proprietários para a necessidade de uma maior vigilância relativamente às formas de interação com os animais, bem como para a adoção de medidas higiénicas cautelares após essa mesma interação.

A power line communication stack for metering, SCADA and large-scale domotic applications

This paper describes the communication stack of the REMPLI system: a structure using power-lines and IPbased networks for communication, for data acquisition and control of energy distribution and consumption. It is furthermore prepared to use alternative communication media like GSM or analog modem connections. The REMPLI system provides communication service for existing applications, namely automated meter reading, energy billing and domotic applications. The communication stack, consisting of physical, network, transport, and application layer is described as well as the communication services provided by the system. We show how the peculiarities of the power-line communication influence the design of the communication stack, by introducing requirements to efficiently use the limited bandwidth, optimize traffic and implement fair use of the communication medium for the extensive communication partners.

Ligand-Induced structural changes in TEM‑1 probed by molecular dynamics and relative binding free energy calculations

The TEM family of enzymes has had a crucial impact on the pharmaceutical industry due to their important role in antibiotic resistance. Even with the latest technologies in structural biology and genomics, no 3D structure of a TEM- 1/antibiotic complex is known previous to acylation. Therefore, the comprehension of their capability in acylate antibiotics is based on the protein macromolecular structure uncomplexed. In this work, molecular docking, molecular dynamic simulations, and relative free energy calculations were applied in order to get a comprehensive and thorough analysis of TEM-1/ampicillin and TEM-1/amoxicillin complexes. We described the complexes and analyzed the effect of ligand binding on the overall structure. We clearly demonstrate that the key residues involved in the stability of the ligand (hot-spots) vary with the nature of the ligand. Structural effects such as (i) the distances between interfacial residues (Ser70−Oγ and Lys73−Nζ, Lys73−Nζ and Ser130−Oγ, and Ser70−Oγ−Ser130−Oγ), (ii) side chain rotamer variation (Tyr105 and Glu240), and (iii) the presence of conserved waters can be also influenced by ligand binding. This study supports the hypothesis that TEM-1 suffers structural modifications upon ligand binding.

Dynamical behaviour of multi-particle large-scale systems

Collective behaviours can be observed in both natural and man-made systems composed of a large number of elemental subsystems. Typically, each elemental subsystem has its own dynamics but, whenever interaction between individuals occurs, the individual behaviours tend to be relaxed, and collective behaviours emerge. In this paper, the collective behaviour of a large-scale system composed of several coupled elemental particles is analysed. The dynamics of the particles are governed by the same type of equations but having different parameter values and initial conditions. Coupling between particles is based on statistical feedback, which means that each particle is affected by the average behaviour of its neighbours. It is shown that the global system may unveil several types of collective behaviours, corresponding to partial synchronisation, characterised by the existence of several clusters of synchronised subsystems, and global synchronisation between particles, where all the elemental particles synchronise completely.

EMMON: a system architecture for large- scale, dense and real-time WSNs

In spite of the significant amount of scientific work in Wireless Sensor Networks (WSNs), there is a clear lack of effective, feasible and usable WSN system architectures that address both functional and non-functional requirements in an integrated fashion. This poster abstract outlines the EMMON system architecture for large-scale, dense, real-time embedded monitoring. EMMON relies on a hierarchical network architecture together with integrated middleware and command&control mechanisms. It has been designed to use standard commercially– available technologies, while maintaining as much flexibility as possible to meet specific applications’ requirements. The EMMON WSN architecture has been validated through extensive simulation and experimental evaluation, including through a 300+ node test-bed, the largest WSN test-bed in Europe to date

Poster: EMMON: a WSN system architecture and toolset for large-scale and dense real-time embedded monitoring

Wireless sensor networks (WSNs) have attracted growing interest in the last decade as an infrastructure to support a diversity of ubiquitous computing and cyber-physical systems. However, most research work has focused on protocols or on specific applications. As a result, there remains a clear lack of effective and usable WSN system architectures that address both functional and non-functional requirements in an integrated fashion. This poster outlines the EMMON system architecture for large-scale, dense, real-time embedded monitoring. It provides a hierarchical communication architecture together with integrated middleware and command and control software. It has been designed to maintain as much as flexibility as possible while meeting specific applications requirements. EMMON has been validated through extensive analytical, simulation and experimental evaluations, including through a 300+ nodes test-bed the largest single-site WSN test-bed in Europe.

The dark side of DEMMON: what is behind the scene in engineering large-scale wireless sensor networks

Most research work on WSNs has focused on protocols or on specific applications. There is a clear lack of easy/ready-to-use WSN technologies and tools for planning, implementing, testing and commissioning WSN systems in an integrated fashion. While there exists a plethora of papers about network planning and deployment methodologies, to the best of our knowledge none of them helps the designer to match coverage requirements with network performance evaluation. In this paper we aim at filling this gap by presenting an unified toolset, i.e., a framework able to provide a global picture of the system, from the network deployment planning to system test and validation. This toolset has been designed to back up the EMMON WSN system architecture for large-scale, dense, real-time embedded monitoring. It includes network deployment planning, worst-case analysis and dimensioning, protocol simulation and automatic remote programming and hardware testing tools. This toolset has been paramount to validate the system architecture through DEMMON1, the first EMMON demonstrator, i.e., a 300+ node test-bed, which is, to the best of our knowledge, the largest single-site WSN test-bed in Europe to date.

EMMON: a WSN system architecture for large scale and dense real-time embedded monitoring

Wireless sensor networks (WSNs) have attracted growing interest in the last decade as an infrastructure to support a diversity of ubiquitous computing and cyber-physical systems. However, most research work has focused on protocols or on specific applications. As a result, there remains a clear lack of effective, feasible and usable system architectures that address both functional and non-functional requirements in an integrated fashion. In this paper, we outline the EMMON system architecture for large-scale, dense, real-time embedded monitoring. EMMON provides a hierarchical communication architecture together with integrated middleware and command and control software. It has been designed to use standard commercially-available technologies, while maintaining as much flexibility as possible to meet specific applications requirements. The EMMON architecture has been validated through extensive simulation and experimental evaluation, including a 300+ node test-bed, which is, to the best of our knowledge, the largest single-site WSN test-bed in Europe to date.

Efficient aggregate computations in large-scale dense WSN

We focus on large-scale and dense deeply embedded systems where, due to the large amount of information generated by all nodes, even simple aggregate computations such as the minimum value (MIN) of the sensor readings become notoriously expensive to obtain. Recent research has exploited a dominance-based medium access control(MAC) protocol, the CAN bus, for computing aggregated quantities in wired systems. For example, MIN can be computed efficiently and an interpolation function which approximates sensor data in an area can be obtained efficiently as well. Dominance-based MAC protocols have recently been proposed for wireless channels and these protocols can be expected to be used for achieving highly scalable aggregate computations in wireless systems. But no experimental demonstration is currently available in the research literature. In this paper, we demonstrate that highly scalable aggregate computations in wireless networks are possible. We do so by (i) building a new wireless hardware platform with appropriate characteristics for making dominance-based MAC protocols efficient, (ii) implementing dominance-based MAC protocols on this platform, (iii) implementing distributed algorithms for aggregate computations (MIN, MAX, Interpolation) using the new implementation of the dominance-based MAC protocol and (iv) performing experiments to prove that such highly scalable aggregate computations in wireless networks are possible.

A synchronous transition protocol with periodicity for global scheduling of multimode real-time systems on multiprocessors

We consider the global scheduling problem of multimode real-time systems upon identical multiprocessor platforms. During the execution of a multimode system, the system can change from one mode to another such that the current task set is replaced with a new task set. Thereby, ensuring that deadlines are met requires not only that a schedulability test is performed on tasks in each mode but also that (i) a protocol for transitioning from one mode to another is specified and (ii) a schedulability test for each transition is performed. In this paper, we extend the synchronous transition protocol SM-MSO in order to take into account mode-independent tasks [1], i.e., tasks of which the execution pattern must not be jeopardized by the mode changes.