Detection and identification of dengue virus isolates from Brazil by a simplified reverse transcription - polymerase chain reaction (RT-PCR) method

We show here a simplified RT-PCR for identification of dengue virus types 1 and 2. Five dengue virus strains, isolated from Brazilian patients, and yellow fever vaccine 17DD as a negative control, were used in this study. C6/36 cells were infected and supernatants were collected after 7 days. The RT-PCR, done in a single reaction vessel, was carried out following a 1/10 dilution of virus in distilled water or in a detergent mixture containing Nonidet P40. The 50 µl assay reaction mixture included 50 pmol of specific primers amplifying a 482 base pair sequence for dengue type 1 and 210 base pair sequence for dengue type 2. In other assays, we used dengue virus consensus primers having maximum sequence similarity to the four serotypes, amplifying a 511 base pair sequence. The reaction mixture also contained 0.1 mM of the four deoxynucleoside triphosphates, 7.5 U of reverse transcriptase, 1U of thermostable Taq DNA polymerase. The mixture was incubated for 5 minutes at 37ºC for reverse transcription followed by 30 cycles of two-step PCR amplification (92ºC for 60 seconds, 53ºC for 60 seconds) with slow temperature increment. The PCR products were subjected to 1.7% agarose gel electrophoresis and visualized by UV light after staining with ethidium bromide solution. Low virus titer around 10 3, 6 TCID50/ml was detected by RT-PCR for dengue type 1. Specific DNA amplification was observed with all the Brazilian dengue strains by using dengue virus consensus primers. As compared to other RT-PCRs, this assay is less laborious, done in a shorter time, and has reduced risk of contamination

Replacement of HIV p24 Ag test by a multiplex RT-PCR method for primary screening of blood donors

Nucleic Acid Testing (NAT) as a tool for primary screening of blood donors became a reality in the end of the 1990 decade. We report here the development of an "in-house" RT-PCR method that allows the simultaneous (multiplex) detection of HCV and HIV-RNA in addition to an artificial RNA employed as an external control. This method detects all HIV group M subtypes, plus group N and O, with a detection threshold of 500 IU/mL. After validation, the method replaced p24 Ag testing, in use for blood donation screening since 1996 at our services. From July 2001 to February 2006, 102,469 donations were tested and 41 (0.04%) were found HIV-RNA reactive. One NAT-only reactive donation (antibody non-reactive) was observed, with subsequent seroconversion of the implied donor, giving a yield of 1:102,469. This rate is in contrast to the international experience that reports a detection of approximately 1:600,000 - 1:3,100,000 of isolated HIV-RNA donations.

Molecular characterization of two rocio flavivirus strains isolated during the encephalitis epidemic in são paulo state, brazil and the development of a one-step rt-pcr assay for diagnosis

Rocio virus (ROCV) was responsible for an explosive encephalitis epidemic in the 1970s affecting about 1,000 residents of 20 coastland counties in São Paulo State, Brazil. ROCV was first isolated in 1975 from the cerebellum of a fatal human case of encephalitis. Clinical manifestations of the illness are similar to those described for St. Louis encephalitis. ROCV shows intense antigenic cross-reactivity with Japanese encephalitis complex (JEC) viruses, particularly with Ilheus (ILHV), St. Louis encephalitis, Murray Valley and West Nile viruses. In this study, we report a specific RT-PCR assay for ROCV diagnosis and the molecular characterization of the SPAn37630 and SPH37623 strains. Partial nucleotide sequences of NS5 and E genes determined from both strains were used in phylogenetic analysis. The results indicated that these strains are closely related to JEC viruses, but forming a distinct subclade together with ILHV, in accordance with results recently reported by Medeiros et al. (2007).

Estimativa da prevalência de infeccções micobacterianas em escolares na Guanabara, através de testes intradérmicos com PPD-Rt 23 E PPD-B

Foram testados 543 escolares do Estado da Guanabara, Brasil, na faixa etária de 12 a 14 anos, quanto à hipersensibilidade tuberculínica ao PPD-Rt 23 e ao PPD-B. Confirmou-se a alta prevalência de sensibilidade às tuberculoproteínas em crianças, obtendo-se 33,65% de reatores fortes ao PPD-Rt 23. Da população estudada 85,63% reagiu ao PPD-B, sendo que 6,26% apresentou reação maior a esta tuberculina que ao PPD-Rt 23, sugerindo uma hipersensibilização pelo bacilo Battey, ou outra microbactéria mais relacionada antigenicamente a esse microrganismo que ao Mycobacterium tuberculosis.

Geração de aplicações a partir de especificações UML como fator de sucesso para o desenvolvimento de sistemas de informação

Dissertação de mestrado integrado em Engenharia e Gestão de Sistemas de Informação

A UML-based approach for modeling industrial control applications

The main purpose of the poster is to present how the Unified Modeling Language (UML) can be used for diagnosing and optimizing real industrial production systems. By using a car radios production line as a case study, the poster shows the modeling process that can be followed during the analysis phase of complex control applications. In order to guarantee the continuity mapping of the models, the authors propose some guidelines to transform the use cases diagrams into a single object diagram, which is the main diagram for the next phases of the development.

Selecting control genes for RT-QPCR using public microarray data.

Background: Gene expression analysis has emerged as a major biological research area, with real-time quantitative reverse transcription PCR (RT-QPCR) being one of the most accurate and widely used techniques for expression profiling of selected genes. In order to obtain results that are comparable across assays, a stable normalization strategy is required. In general, the normalization of PCR measurements between different samples uses one to several control genes (e. g. housekeeping genes), from which a baseline reference level is constructed. Thus, the choice of the control genes is of utmost importance, yet there is not a generally accepted standard technique for screening a large number of candidates and identifying the best ones. Results: We propose a novel approach for scoring and ranking candidate genes for their suitability as control genes. Our approach relies on publicly available microarray data and allows the combination of multiple data sets originating from different platforms and/or representing different pathologies. The use of microarray data allows the screening of tens of thousands of genes, producing very comprehensive lists of candidates. We also provide two lists of candidate control genes: one which is breast cancer-specific and one with more general applicability. Two genes from the breast cancer list which had not been previously used as control genes are identified and validated by RT-QPCR. Open source R functions are available at http://www.isrec.isb-sib.ch/similar to vpopovic/research/ Conclusion: We proposed a new method for identifying candidate control genes for RT-QPCR which was able to rank thousands of genes according to some predefined suitability criteria and we applied it to the case of breast cancer. We also empirically showed that translating the results from microarray to PCR platform was achievable.

Fidelidad al modelo especificado en el código generado automáticamente por la herramienta CASE Poseidon for UML

L'objectiu principal d'aquest treball és estudiar el grau de fidelitat al model especificat que manté el codi generat automàticament per l'eina CASE Poseidon Professional for UML. Concretament, estudiarem el codi que es genera en Java i SQL mitjançant un diagrama de classes determinat.

Intérêt des faibles doses de rt-PA dans les syndromes inflammatoires et hémorragiques du segment antérieur: étude clinique et revue de la littérature [Treatment of anterior segment fibrinous reactions and hemorrhage with intracameral low dose rt-PA: clinical study and review of the literature].

PURPOSE: to evaluate the efficacy and the safety of low dose intraocular tissue plasminogen activator (rt-PA) in the treatment of traumatic hyphema and postoperative fibrinous membrane. METHODS: Six microg to 10 microg of rt-PA was injected into the anterior chamber to treat severe fibrinous postoperative membranes and total traumatic hyphemae. RESULTS: Thirteen eyes of 13 patients were treated. Four cases of traumatic hyphema and 9 cases of fibrinous membranes were included. Complete fibrinolysis within 24 hours was observed in 4 cases (30.8%). A partial success was noted in 7 eyes (53.8%). No beneficial effect was observed in two cases of traumatic hyphema associated with intravitreal hemorrhage after penetrating trauma. No side effect attributable to rt-PA occurred. CONCLUSION: Low dose intraocular rt-PA appears to be safe and effective in the treatment of postoperative fibrinous membrane and endocular hemorrhage limited to the anterior chamber.

Comparació de Diagrames UML

El nucli del projecte consisteix en desenvolupar una aplicació permeti determinar si dos models UML són iguals i , en cas de no ser-ho, determinar quines són les diferències entre ells. Donada la gran varietat de models UML, el projecte es centra en els diagrames de classes.

Aplicativo web para la corrección de modelos UML/OCL

Introducció d'un entorn virtual d' autoaprenentatge que permeti als estudiants millorar les seves habilitats de modelització, una peça clau en la seva capacitació com a professionals de la informàtica adaptats a les demandes de la societat actual.

Creació de taules d'una base de dades relacional a partir del diagrama de classes d'anàlisi d'UML

És ben sabut que les úniques etapes del cicle de vida del programari que necessàriament s'han d'especificar són les de recollida de requisits i l'anàlisi o l'especificació del programari. La resta (disseny, implementació i prova) es pot generar d'una manera més o menys automàtica a partir de l'anàlisi. En aquest PFC hem volgut estudiar la viabilitat de la construcció automàtica de codi SQL a partir de diagrames de classes d'anàlisi UML. S'ha estès l'eina de modelatge UML Poseidon amb un connector, de manera que amb una interfície molt simple es pot obtenir molt ràpidament l'esquema bàsic d'una base de dades, incloent-hi les taules, les seves columnes, claus rimàries i foranes, i també els disparadors (i les claus úniques) necessaris per a garantir les restriccions de cardinalitat de les associacions.