Neurotoxic activity and ultrastructural changes in muscles caused by the brown widow spider Latrodectus geometricus venom

Brown widow spider (Latrodectus geometricus) venom (BrWSV) produces few local lesions and intense systemic reactions such as cramps, harsh muscle pains, nausea, vomiting and hypertension. Approximately 16 protein bands under reducing conditions and ~ 14 bands under non-reducing conditions on a 12.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis were observed. Neurotoxic clinical manifestations were confirmed in vivo, while proteolytic activity was demonstrated on gelatine film. Severe ultrastructural damages in mice skeletal muscles were observed at 3, 6, 12 and 24 h postinjection with at total of 45 µg of venom protein. Infiltration of eosinophils and ruptures of the cellular membranes were observed in the muscles along with swelling of the nuclear cover and interruption of the collagen periodicity. Altered mitochondrias and autophage vacuoles, nuclear indentation and mitochondria without cristae, slight increment of intermyofibrillar and subsarcolemic spaces and myelinic figures formation were also observed. In the capillary, endothelial membrane unfolding into the lumen was noticed; along with myelinic figures compatible with a toxic myopathy. Swollen sarcotubular systems with lysis of membrane, intense mitochondria autophagia and areas without pinocytic vesicles were observed. Swollen mitochondria surrounded by necrotic areas, myofibrillar disorganization and big vacuolas of the sarcotubular system, degenerated mitochondrium with formation of myelinic figure was seen. Glycogenosomes with small particulate, muscle type glycogen was noticed. Autophagic vacuole (autophagolysosomes) and necrotic areas were also noticed. These damages may be due to interactive effects of the multifactorial action of venom components. However, Latrodectus geometricus venom molecules may also be utilized as neuro therapeutic tools, as they affect neuronal activities with high affinity and selectivity. To our knowledge, the present study is the first ultrastructural report in the literature of muscle injuries and neurological and proteolytic activities caused by BrWSV.

Extended-spectrum beta-lactamases among Enterobacteriaceae isolated in a public hospital in Brazil

Extended-spectrum β-lactamases (ESBL) in enterobacteria are recognized worldwide as a great hospital problem. In this study, 127 ESBL-producing Enterobacteriaceae isolated in one year from inpatients and outpatients at a public teaching hospital at São Paulo, Brazil, were submitted to analysis by PCR with specific primers for blaSHV, blaTEM and blaCTX-M genes. From the 127 isolates, 96 (75.6%) Klebsiella pneumoniae, 12 (9.3%) Escherichia coli, 8 (6.2%) Morganella morganii, 3 (2.3%) Proteus mirabilis, 2 (1.6%) Klebsiella oxytoca, 2 (1.6%) Providencia rettgeri, 2 (1.6%) Providencia stuartti, 1 (0.8%) Enterobacter aerogenes and 1 (0.8%) Enterobacter cloacae were identified as ESBL producers. BlaSHV, blaTEM and blaCTX-M were detected in 63%, 17.3% and 33.9% strains, respectively. Pulsed field gel eletrophoresis genotyping of K. pneumoniae revealed four main molecular patterns and 29 unrelated profiles. PCR results showed a high variety of ESBL groups among strains, in nine different species. The results suggest the spread of resistance genes among genetically different strains of ESBL-producing K. pneumoniae in some hospital wards, and also that some strongly related strains were identified in different hospital wards, suggesting clonal spread in the institutional environment.

Vólvulo Intestinal en el Periodo Neonatal: 8 Años de Experiencia en un Hospital Pediátrico Terciario

Análise dos casos de volvo intestinal ocorridos no período neonatal nos últimos 8 anos (2002 a 2010). Material e métodos: Foram estudados os recém-nascidos admitidos na UCIN cujo diagnóstico de saída foi volvo intestinal. Foram estudados os seguintes parâmetros: idade gestacional e pós-natal, apresentação clínica e imagiológica, intervenção cirúrgica e resultados. Resultados: Foram identificados 15 doentes 7 dos quais no último ano do estudo. Sete RN eram pré-termo (PT) ou ex pré-termo. A mediana de peso ao nascer foi de 2665g (660-3900); 4 RN eram muito baixo peso. A mediana de idade de início dos sintomas foi 7 dias; em 5 RN a doença teve início nas primeiras 24 horas de vida; em 3 destes, o volvo ocorreu in utero. Sinais e sintomas: grande distensão abdominal-12; resíduo gástrico bilioso-11; alterações da parede abdominal-5; dejecções com sangue-4; instabilidade hemodinâmica-6. Imagiologia: grande distensão de ansas, sem ar ectópico- 10 doentes; ausência de ar no abdómen-4; trânsito intestinal contrastado sugestivo de malrotação e volvo-3; ecografia e Doppler abdominal com sinal de “whirlpool”-2. Todos foram submetidos a cirurgia de urgência, sendo o volvo confirmado intraoperatoriamente; foi necessária ressecção intestinal em 9 doentes; 3 ficaram com síndrome do intestino curto; registou-se um óbito por falência multi-orgânica no período pós-operatório. Conclusão: Foi encontrado um elevado número de casos de volvo intestinal em RN pré-termo ou ex pré-termo, de volvo in utero e de elevada ocorrência de casos no último ano do estudo. Resíduo gástrico bilioso e distensão abdominal foram os sintomas mais frequentes de volvo e devem ser tomados em consideração no diagnóstico diferencial com outras situações cirúrgicas abdominais. As sequelas são potencialmente graves.

Décroissance volontaire et perspectives en matière d'alimentaire

Dissertação apresentada para cumprimento dos requisitos necessários à obtenção do grau de Mestre em Ecologia Humana e Problemas Sociais Contemporâneos

Estudio de la gestión de la variable ambiental en la generación termoeléctrica

En la actualidad, el cambio climático es uno de los temas de mayor preocupación para la población mundial y los científicos de todo el mundo. Debido al crecimiento de la población de forma exponencial, la demanda de energía aumenta acorde con ello, por lo que las actividades de producción energética aumentan consecuentemente, siendo éstas las principales causantes de la aceleración del cambio climático. Pese a que muchos países previamente habían apostado por la producción energética mediante tecnologías limpias a partir de energías renovables, hoy en día es imposible prescindir de los combustibles fósiles pues, junto a la energía nuclear, suponen el mayor porcentaje dentro del mix energético de los países más grandes del mundo, por lo que el cambio debe ser global y con todos los países implicados al unísono. Por ello, los países desarrollados decidieron acordar una serie de leyes y normas para la regulación y el control de la expansión energética en el mundo, mediante programas de incentivo a las empresas para la producción de energía limpia, libre de emisiones, sustituyendo y mejorando los procesos tecnológicos para que garanticen un desarrollo sostenible. De esta forma, se conseguiría también reducir la dependencia energética de los países productores de los recursos fósiles más importantes y a su vez, ayudar a otros sectores a diversificar su negocio y mejorar así la economía de las áreas colindantes a las centrales de producción térmica. Gracias a estos programas de incentivo o, también llamados mecanismos de flexibilidad, las empresas productoras de energía, al acometer inversiones en tecnologia limpia, dejan de emitir gases de efecto invernadero a la atmósfera. Por tanto, gracias al comercio de emisiones y al mercado voluntario, las empresas pueden vender dichas emisiones aumentando la rentabilidad de sus proyectos, haciendo más atractivo de por sí el hecho de invertir en tecnología limpia. En el proyecto desarrollado, se podrá comprobar de una forma más extensa todo lo anteriormente citado. Para ello, se desarrollará una herramienta de cálculo que nos permitirá analizar los beneficios obtenidos por la sustitución de un combustible fósil, no renovable, por otro renovable y sostenible, como es la biomasa. En esta herramienta se calcularán, de forma estimada, las reducciones de las emisiones de CO2 que supone dicha sustitución y se hallará, en función del valor de las cotizaciones de los bonos de carbono en los diferentes mercados, cuál será el beneficio económico obtenido por la venta de las emisiones no emitidas que supone esta sustitución. Por último, dicho beneficio será insertado en un balance económico de la central donde se tendrán en cuenta otras variables como el precio del combustible o las fluctuaciones del precio de la electricidad, para hallar finalmente la rentabilidad que supondría la inversión de esta adaptación en la central. Con el fin de complementar y aplicar la herramienta de cálculo, se analizarán dos casos prácticos de una central de carbón, en los cuales se decide su suscripción dentro del contexto de los mecanismos de flexibilidad creados en los acuerdos internacionales.

Regionalización del Cuidado Perinatal en Portugal - Resultados

Lo Servicio Nacional de Salud en Portugal fue criado en 1979, universal e gratis. Las primeras UCIN surgieran en 1980, en 1985 fue criada la Sociedad Portuguesa de Neonatología, en 1987 lo sistema de transporte neonatal, en mismo año fue nominada una Comisión de Peritos en Perinatología. En 1989 fue nominado el Comité Nacional de la Mujer y del niño y empezó la Reforma de los cuidados de salud perinatal. Era un programa de 9 años en etapas de 3 años que incluía el ccierre de Hospitales con menos de 1500 partos/año, categorización de los hospitales en niveles de cuidados e la creación de Unidades Coordinadoras entre Centros de Salud y Hospitales. Las UCIN y Intermedios neonatales fueran equipados y definido el número necesario de obstetras, pediatras y enfermeras e fue hecha formación en Cuidados Intensivos Neonatales. Los Centros de Salud no tienen partos e controlan el embarazo normal; los hospitales Nivel I no tienen partos; los de Nivel II tienen partos normales y de bajo riesgo, por lo menos 1500/año, obstetras, Unidad de Cuidados Intermedios, Pediatras con formación en neonatología, ventilación por períodos cortos. Los Hospitales de Nivel III tienen partos de bajo y alto riesgo, obstetras y neonatólogos, UCIN, formación en obstetricia y neonatología e investigación. Las UCIN tienen ventilación de longa duración, nutrición parenteral, cuidados de recién nacidos con menos de 1500g, condiciones quirúrgicas, son centros de enseñanza e investigación. Deben tener neonatólogos y pediatras con competencia en neonatología 24h por día, 1.5 camas /1000 partos, 1 enfermera para 2 logares de intensivos – 2,5 enfermeras por cada cama de intensivos - deben estar localizadas en maternidades con >3000 partos. Hay también reglas para translado para nível III pré-natal y pós-natal. Lo impacto fue que la tasa de partos hospitalares aumentó hasta 99% la mortalidad fetal e fetal tardia disminuiu hasta 3,7 e 2,5/1000 NV+MN, la mortalidad perinatal con más de 28 semanas para 4,6/1000 NV+ MN, la neonatal para 2,4/1000NV e la infantil para 3,6. Las mejores condiciones socioeconómicas y nivel de educación pueden justificar parte de estos resultados pero la organización de los cuidado perinatales fue ciertamente una grande razón de mejoría.

Neutralization of bitis parviocula (Ethiopian mountain adder) venom by the south african institute of medical research (SAIMR) antivenom

BACKGROUND: The Ethiopian mountain adder (Bitis parviocula) is a viperid known only from a few locations in southwestern Ethiopia. METHODS: a total of 30 µg of B. arietans and B. parviocula venoms were run on a 10-20% Tricine gel. To assay lethality dose fifty (LD50), five groups of eight mice for each venom were used. Hemorrhagic activity for crude venom was tested. Fibrinogenolytic activity of crude venom was measured using (2.5 mg/mL) of fibrinogen solution and (0.03 mg/mL) of crude venom. Gelatinase activity of the venom was tested on a Kodak X-OMAT TM film. Crude venoms of B. parviocula and B. arietans were tested for their abilities to affect clotting time, clotting rate and platelet function on whole human blood. RESULTS: The (SAIMR) antivenom was confirmed in this study to neutralize the lethal activity of venom from Bitis parviocula. The ED50s of SAIMR antivenom on B. parviocula and B. arietans neutralized half of 18.2 and 66.7 mg of venom, respectively. The hemorrhagic activities (MHDs) of B. parviocula and B. arietans were 0.88 and 1.7 µg, respectively. Bitis arietans and B. parviocula venoms degradated α and β chains at different times. The γ chains remained unaffected. Bitis parviocula venom did not exhibit gelatinase activity, while B. arietans had a MGD of 6.9 µg. At 3 mg/mL, the crude venoms of B. parviocula and B. arietans did not significantly affect clotting time or clotting rate. CONCLUSIONS: The SAIMR antivenom is very effective in neutralizing the venom of B. parviocula and should be considered in treating envenomations by these snakes.

Potential utility of hyperbaric oxygen therapy and propolis in enhancing the leishmanicidal activity of glucantime

In this study we investigated the efficacy of hyperbaric oxygen (HBO) therapy, alone or combined with the pentavalent antimonial glucantime on Leishmania amazonensis infection. In parallel, the effect of Brazilian red propolis gel (propain) alone or combined with glucantime on L. amazonensis infection was evaluated. The inhibition of the infection in macrophages treated with glucantime in combination with HBO exposition was greater than that of macrophages treated with glucantime alone or HBO alone. The susceptible mouse strain BALB/c infected in the shaved rump with L. amazonensis treated with glucantime and exposed to HBO showed: time points in the course of the disease in which lesions were smaller than those of mice treated with glucantime alone and revascularization of the skin in the lesion site; interferon-gamma (IFN-g) levels were not elevated in lymph node cells from these animals. Propain alone was not efficient against lesions, although less exudative lesions were observed in animals treated with propain alone or combined with glucantime. These results reveal the potential value of HBO and red propolis in combination with glucantime for treating cutaneous leishmaniasis and encourage further studies on the effect of more aggressive HBO, propolis and glucantime therapies on different mouse models of leishmaniasis.

One Anastomosis Gastric Bypass versus Roux en Y Gastric Bypass as Salvage Technique after Failed Gastric Band: a Retrospective Analysis of 80 Cases

The classical “Roux en Y Gastric Bypass” (RYGBP) is still the standard technique between all the ones being used nowadays. The “One anastomosis gastric bypass” (OAGBP), is an evolution of the “Minigastic bypass” described by Robert Rutledge in 2001, is a well known and progressively frequent but still controversial technique. In our group, after an experience of 10 years using the RYGBP as a salvage surgery after failed gastric banding, in 132 cases, we decided to adopt the OAGBP as our preferential bariatric technique also in this situation. The theoretical main reasons for that shift are related to the increased safety, maximized weight loss, long term weight loss maintenance and reversibility of the operation. Method: Retrospectively we evaluated data of the surgical management of revisional cases for conversion, after failed or complicated gastric bands to gastric bypass. We selected the last 40 cases of each technique since May 2010. Results: All cases were performed by laparoscopy without any conversion. In both groups the conversion has been performed in one single step (17 cases, 42,5%). Data showed lower morbidity with OAGBP (2,5% against 7,5%) and better weight loss in theOAGBP cohort after a median follow up of 16months (67%against 55%) in patients revised after gastric band failure or complications. None had statistic significance (p>0,1) by the chi-square contingency table analysis.Conclusion: It seems to there is a difference in favour of OAGBP for conversion of complicated gastric bands. In this study we didn’t found statistic significance probably because of the short numbers. Prospective and more powerful studies are necessary to evaluate the benefit of the studied procedure.

APPLICATION OF 6-NITROCOUMARIN AS A SUBSTRATE FOR THE FLUORESCENT DETECTION OF NITROREDUCTASE ACTIVITY IN Sporothrix schenckii

Introduction Sporothrix schenckii is a thermal dimorphic pathogenic fungus causing a subcutaneous mycosis, sporotrichosis. Nitrocoumarin represents a fluorogenic substrate class where the microbial nitroreductase activity produces several derivatives, already used in several other enzyme assays. The objective of this study was the analysis of 6-nitrocoumarin (6-NC) as a substrate to study the nitroreductase activity in Sporothrix schenckii. Methods Thirty-five samples of S. schenckii were cultivated for seven, 14 and 21 days at 35 °C in a microculture containing 6-nitrocoumarin or 6-aminocoumarin (6-AC) dissolved in dimethyl sulfoxide or dimethyl sulfoxide as a negative control, for posterior examination under an epifluorescence microscope. The organic layer of the seven, 14 and 21-day cultures was analyzed by means of direct illumination with 365 nm UV light and by means of elution on G silica gel plate with hexane:ethyl acetate 1:4 unveiled with UV light. Results All of the strains showed the presence of 6-AC (yellow fluorescence) and 6-hydroxylaminocoumarin (blue fluorescence) in thin layer chromatography, which explains the green fluorescence observed in the fungus structure. Conclusion The nitroreductase activity is widely distributed in the S. schenckii complex and 6-NC is a fluorogenic substrate of easy access and applicability for the nitroreductase activity detection.

Implementação do referencial IFS alimentar numa indústria de produtos pré-confeccionados e ultracongelados integração com a NP EN ISO 9001:2008

Dissertação para a obtenção do Grau de Mestre em Tecnologia e Segurança Alimentar – Qualidade Alimentar

Smqnr VARIANTS IN CLINICAL ISOLATES OF Stenotrophomonas maltophilia IN BRAZIL

SUMMARYStenotrophomonas maltophilia contains a novel chromosomally-encoded qnr gene named Smqnr that contributes to low intrinsic resistance to quinolone. We described Smqnr in 13 clinical isolates of S. maltophilia from two Brazilian hospitals, over a 2-year period. The strains were identified by API 20 NE (bioMérieux, France). Susceptibility by microdilution method to trimetroprim/sulfamethoxazole, ciprofloxacin, levofloxacin, minocycline, ceftazidime, chloramphenicol and ticarcillin/clavulanate was performed according to CLSI. PCR detection of Smqnr gene was carried out. The sequence of Smqnr was compared with those deposited in GenBank. Pulsed-field gel electrophoresis (PFGE) of all strains was performed. Thirteen Smqnr positives isolates were sequenced and three novel variants of Smqnr were identified. All 13 Smqnr isolates had distinguishable patterns by PFGE. This is the first report of Smqnr in S. maltophilia isolated in Brazil.

EVALUATION OF FOUR DIFFERENT DNA EXTRACTION METHODS IN COAGULASE-NEGATIVE STAPHYLOCOCCI CLINICAL ISOLATES

Currently there are several methods to extract bacterial DNA based on different principles. However, the amount and the quality of the DNA obtained by each one of those methods is highly variable and microorganism dependent, as illustrated by coagulase-negative staphylococci (CoNS) which have a thick cell wall that is difficult to lyse. This study was designed to compare the quality and the amount of CoNS DNA, extracted by four different techniques: two in-house protocols and two commercial kits. DNA amount and quality determination was performed through spectrophotometry. The extracted DNA was also analyzed using agarose gel electrophoresis and by PCR. 267 isolates of CoNS were used in this study. The column method and thermal lyses showed better results with regard to DNA quality (mean ratio of A260/280 = 1.95) and average concentration of DNA (), respectively. All four methods tested provided appropriate DNA for PCR amplification, but with different yields. DNA quality is important since it allows the application of a large number of molecular biology techniques, and also it's storage for a longer period of time. In this sense the extraction method based on an extraction column presented the best results for CoNS.

Candida albicans PROTEIN PROFILE CHANGES IN RESPONSE TO THE BUTANOLIC EXTRACT OF Sapindus saponariaL.

Candida albicans is an opportunistic human pathogen that is capable of causing superficial and systemic infections in immunocompromised patients. Extracts of Sapindus saponaria have been used as antimicrobial agents against various organisms. In the present study, we used a combination of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify the changes in protein abundance of C. albicans after exposure to the minimal inhibitory concentration (MIC) and sub-minimal inhibitory concentration (sub-MIC) of the butanolic extract (BUTE) of S. saponaria and also to fluconazole. A total of six different proteins with greater than 1.5 fold induction or repression relative to the untreated control cells were identified among the three treatments. In general, proteins/enzymes involved with the glycolysis (GPM1, ENO1, FBA1), amino acid metabolism (ILV5, PDC11) and protein synthesis (ASC1) pathways were detected. In conclusion, our findings reveal antifungal-induced changes in protein abundance of C. albicans. By using the previously identified components of the BUTE of S. saponaria(e.g., saponins and sesquiterpene oligoglycosides), it will be possible to compare the behavior of compounds with unknown mechanisms of action, and this knowledge will help to focus the subsequent biochemical work aimed at defining the effects of these compounds.

Prácticas de Prescripción de Nutrición Parenteral Neonatal en Portugal

Introducción: La utilización de normas de orientación para la nutrición parenteral neonatal mejora la eficiencia y la seguridad de su prescripción. Objetivo: Evaluar la práctica de prescripción de nutrición parenteral neonatal en Portugal y el cumplimiento del Consenso Nacional (2008). Métodos: Encuesta de respuesta múltiple sobre la prescripción de nutrición parenteral (NP) enviada a los coordinadores de las 50 unidades de cuidados especiales neonatales portuguesas, tanto públicas como privadas, siendo 25 de nível ii y 25 de nivel iii. Resultados: Se prescribe NP en 32 unidades neonatales, habiendo 23 (71,9%) respondido la encuesta. De estas, 19 (82,6%) afirman tener como referencia el Consenso Nacional y las restantes siguen protocolos internos; 17 (73,9%) afirman preferir la prescripción mediante soporte informático. En recién nacidos pretérmino, la mayoría reporta una administración cautelosa de líquidos en la primera semana posnatal; inicio de aminoácidos desde el primer día posnatal con 1,5-3 g/kg/día y aumento hasta 3-4 g/kg/día; inicio de lípidos en los 3 días posnatales com 1 g/kg/día y aumento hasta 3 g/kg/día; administración de 40-70 mg/kg/día de calcio y fósforo, con un ratio calcio:fósforo de 1,7:1 (mg:mg), y estimación de la osmolaridad de las soluciones y control semanal de la trigliceridemia, uremia, fosforemia y función hepática. Conclusiones: Por la elevada tasa de respuesta la muestra probablemente es representativa de la práctica de prescripción de nutrición parenteral neonatal en Portugal. La mayoría de las unidades tienen como referencia el Consenso Nacional, lo que contribuye a unos mejores cuidados de los recién nacidos.