Caracterização de cepas de referência de Leptospira sp utilizando a técnica de pulsed field gel electrophoresis

INTRODUÇÃO: A leptospirose é uma zoonose endêmica, mundialmente distribuída, causada por bactérias do gênero Leptospira. Este gênero compreende espécies patogênicas e saprofíticas, com mais de 200 sorovares distintos, dificultando sua caracterização. A técnica de pulsed field gel electrophoresis tem sido empregada como uma ferramenta para auxiliar nesta caracterização. Os objetivos deste trabalho foram padronizar a técnica de PFGE, determinar os perfis moleculares das cepas de referência utilizadas pelo Laboratório de Referência Nacional para Leptospirose/Centro Colaborador da Organização Mundial de Saúde para Leptospirose e criar um banco de dados com estes perfis. MÉTODOS: Foram analisadas, por PFGE, dezenove cepas utilizando a enzima de restrição NotI. RESULTADOS: Cada cepa apresentou um perfil único que pode ser considerado como uma identidade genômica específica, com exceção dos sorovares Icterohaemorrhagiae e Copenhageni, cujos perfis foram indistinguíveis. CONCLUSÕES: Dessa forma, foi possível a criação de um banco de perfis moleculares que está sendo utilizado no Laboratório para a comparação e identificação de cepas isoladas de quadros clínicos.

Morphological investigation of Toxoplasma gondii in vivo by a multiple beam interference microscope

A recently developed technique, namely multiple beam interference microscopy, has been applied to investigate the morphology of the parasite Toxoplasma gondii for the first time. The interference pattern obtained from the multiple internal reflection of a T. gondii, sandwiched between a glass plate and a cover plate, was focused on the objective of a conventional microscope. Because of the enhance contrast, several details of sub cellular structure and separating compartments are clearly visible. Details reveal the presence of a nucleus, lipid body, dense granule, rhoptry and amylopectin. The wall thickness of the membrane of the lipid body and the amylopectin is of the order of 0.02 µm and can be clearly distinguished with the help of the present technique. The same parasite has also been examined with the help of atomic force microscopy, and because of its thick membrane, the inner structural details were not observed at all. Sub cellular details of T. gondii observed with the present technique have been reported earlier only by low amplification transmission electron microscopy and not by any optical microscopic technique.

Pulsed-field gel electrophoresis, virulence determinants and antimicrobial susceptibility profiles of type Ia group B streptococci isolated from humans in Brazil

Group B streptococci (GBS) infections occur worldwide. Although serotyping has been used for epidemiologic purposes, this does not accurately characterize enough members of a genetically heterogeneous bacterial population. The aims of this work were to evaluate the genetic diversity of 45 type Ia GBS strains isolated in Brazil by pulsed-field gel electrophoresis as well as to evaluate antimicrobial susceptibility profiles and identify virulence genes. Twenty-four strains were assigned to cluster A. All strains under study contained the hylB and scpB genes. The bca gene was detected in only 10 strains and none of the streptococci carried the bac gene. Thirty-nine strains were resistant to tetracycline.

Time-based distribution of Staphylococcus saprophyticus pulsed field gel-electrophoresis clusters in community-acquired urinary tract infections

The epidemiology of urinary tract infections (UTI) by Staphylococcus saprophyticus has not been fully characterised and strain typing methods have not been validated for this agent. To evaluate whether epidemiological relationships exist between clusters of pulsed field gel-electrophoresis (PFGE) genotypes of S. saprophyticus from community-acquired UTI, a cross-sectional surveillance study was conducted in the city of Rio de Janeiro, Brazil. In total, 32 (16%) female patients attending two walk-in clinics were culture-positive for S. saprophyticus. Five PFGE clusters were defined and evaluated against epidemiological data. The PFGE clusters were grouped in time, suggesting the existence of community point sources of S. saprophyticus. From these point sources, S. saprophyticus strains may spread among individuals.

Utilização de uma interface do tipo "particle beam" para a obtenção de espectros de massas de compostos pouco voláteis em solução

Due to source contamination and wearing of instrument components problems caused by the direct insertion probe technique, a new way of introduction of low volatile compounds into mass spectrometer was tested. This new scheme comprises the introduction of the low volatile compounds solutions via a six port valve connected to a particle beam interface. Solutions of isatin were injected into this system and the best results were obtained with CH2Cl2, CH3OH and CH3CN. The solution inlet system has shown to be advantageous over the conventional way of direct insertion probe introduction.

Irradiação de polímeros com feixe de elétrons: caracterização de íons positivos através da técnica de tempo-de-vôo

With the aim of studying the interaction of fast electrons with solid surfaces we have developed an experimental set-up based on electron stimulated desorption (ESD) coupled to time-of-flight (TOF) mass spectrometry. Poly(methyl methacrylate) and poly(vynil chloride) samples have been irradiated by a pulsed electron beam of 1.2 keV and 0.18 µs FWHM. The results show that H+ is the main ionic species to desorb after electron bombardment. In addition, other ionic fragments were also observed and assigned. These results show the potentiality of this technique in the study of ESD of polymers.

LOW COST ANALYZER FOR THE DETERMINATION OF PHOSPHORUS BASED ON OPEN-SOURCE HARDWARE AND PULSED FLOWS

The need for automated analyzers for industrial and environmental samples has triggered the research for new and cost-effective strategies of automation and control of analytical systems. The widespread availability of open-source hardware together with novel analytical methods based on pulsed flows have opened the possibility of implementing standalone automated analytical systems at low cost. Among the areas that can benefit from this approach are the analysis of industrial products and effluents and environmental analysis. In this work, a multi-pumping flow system is proposed for the determination of phosphorus in effluents and polluted water samples. The system employs photometric detection based on the formation of molybdovanadophosphoric acid, and the fluidic circuit is built using three solenoid micropumps. The detection is implemented with a low cost LED-photodiode photometric detection system and the whole system is controlled by an open-source Arduino Uno microcontroller board. The optimization of the timing to ensure the color development and the pumping cycle is discussed for the proposed implementation. Experimental results to evaluate the system behavior are presented verifying a linear relationship between the relative absorbance and the phosphorus concentrations for levels as high as 50 mg L-1.

Evaluation of left ventricular diastolic echocardiographic parameters in healthy dogs by pulsed-wave Doppler

Left ventricular diastolic dysfunction plays an important role on heart failure progression. In order to obtain additional reference values of left ventricular diastolic parameters and investigate influence of common variables, peak E wave (peak E), peak A wave (peak A), E/A ratio (E/A), E wave deceleration time (EDT) and isovolumic relaxation time (IRVT) were studied in 40 clinically healthy dogs, by pulsed wave Doppler. The following values were obtained: peak E = 0.747 ± 0.117 m/s, peak A = 0.487 ± 0.062 m/s, E/A = 1.533 ± 0.198, EDT = 88.7 ± 9.2 ms and IRVT = 0.080 ± 0.009 s. Some parameters were influenced by heart rate (peak E, peak A and IRVT), by age (peak A and E/A) and by body weight (TRIV). Gender influence was absent. Values obtained can be used as reference for canine specimens but its interpretation should consider on the influence of related variables.

The pulsed wire anemometer: review and further developments

The purpose of the present paper is to review work that has been done on the pulsed wire anemometer technique and also suggest further developments that could be made in its range of application. The aper discusses the three types of probes that have been used in pulsed wire anemometry: the crossed wire velocity probe, the parallel wire wall shear stress probe and the parallel wire velocity probe. The work shows that the crossed wire and the parallel wire techniques can be used to make velocity, turbulence and wall shear stress measurements in highly turbulent flows without any upper restriction on turbulence level. Comments are also made on the potential of a parallel wire probe for use in highly turbulent flows that would enable higher order velocity cross-product terms to be measured.

Chaos and Unpredictability in the Vibration of an Elasto-Plastic Beam

This contribution discusses the nonlinear dynamics of a pin-ended elasto-plastic beam with both kinematic and isotropic hardening. An iterative numerical procedure based on the operator split technique is developed in order to deal with the nonlinearities in the equations of motion. Free and forced responses for harmonic sinusoidal and square wave excitations are investigated. Numerical simulations present many interesting behaviors such as jump phenomena, sensitivity to initial conditions, chaos and transient chaos. These results indicate that there are practical problems in predicting the response of the beam even when periodic steady state response is expected.

Experimental Vibration Characteristics of a Beam with Nonlinear Support Using Receptance Coupling Analysis

This paper applies the Multi-Harmonic Nonlinear Receptance Coupling Approach (MUHANORCA) (Ferreira 1998) to evaluate the frequency response characteristics of a beam which is clamped at one end and supported at the other end by a nonlinear cubic stiffness joint. In order to apply the substructure coupling technique, the problem was characterised by coupling a clamped linear beam with a nonlinear cubic stiffness joint. The experimental results were obtained by a sinusoidal excitation with a special force control algorithm where the level of the fundamental force is kept constant and the level of the harmonics is kept zero for all the frequencies measured.

Typing of Enterococcus faecium by polymerase chain reaction and pulsed field gel electrophoresis

Polymerase chain reaction (PCR) with JB1 or REP consensus oligonucleotides and pulsed field gel electrophoresis (PFGE) were used to study genomic DNA extracted from 31 strains of enterococci. Eleven ATCC strains, representative of 11 species of Enterococcus, were initially tested by JB1-PCR, revealing that Enterococcus malodoratus and Enterococcus hirae presented identical banding patterns. Eight Enterococcus faecium isolates from Stanford University and 12 from São Paulo Hospital were studied by JB1-PCR, REP-PCR 1/2R and PFGE. Among the isolates from Stanford University, 5 genotypes were defined by JB1-PCR, 7 by REP-PCR 1/2R and 4 by PFGE. Among the isolates from São Paulo Hospital, 9 genotypes were identified by JB1-PCR, 6 by REP-PCR and 5 by PFGE. The three methods identified identical genotypes, but there was not complete agreement among them.

Pulsed ultrasound therapy accelerates the recovery of skeletal muscle damage induced by Bothrops jararacussu venom

We studied the effect of pulsed ultrasound therapy (UST) and antibothropic polyvalent antivenom (PAV) on the regeneration of mouse extensor digitorum longus muscle following damage by Bothrops jararacussu venom. Animals (Swiss male and female mice weighing 25.0 ± 5.0 g; 5 animals per group) received a perimuscular injection of venom (1 mg/kg) and treatment with UST was started 1 h later (1 min/day, 3 MHz, 0.3 W/cm², pulsed mode). Three and 28 days after injection, muscles were dissected and processed for light microscopy. The venom caused complete degeneration of muscle fibers. UST alone and combined with PAV (1.0 mL/kg) partially protected these fibers, whereas muscles receiving no treatment showed disorganized fascicules and fibers with reduced diameter. Treatment with UST and PAV decreased the effects of the venom on creatine kinase content and motor activity (approximately 75 and 48%, respectively). Sonication of the venom solution immediately before application decreased the in vivo and ex vivo myotoxic activities (approximately 60 and 50%, respectively). The present data show that UST counteracts some effects of B. jararacussu venom, causing structural and functional improvement of the regenerated muscle after venom injury.