Proteómica microbiana: análise comparativa da bactéria redutora de sulfato Desulfovibrio desulfuricans ATCC 27774 e das actinobactérias Salinispora arenicola e Salinispora pacifica

As bactérias redutoras de sulfato (BRS) são um grupo diversificado de micro-organismos anaeróbios que obtêm energia a partir da redução dissimilativa do sulfato. Algumas espécies de BRS possuem versatilidade a nível respiratório, como é o caso de Desulfovibrio desulfuricans ATCC 27774, devido à utilização de aceitadores finais de eletrões alternativos. Neste contexto, o objetivo da primeira parte desta dissertação consistiu na identificação das ferramentas metabólicas (proteínas) envolvidas na flexibilidade respiratória desta bacteria induzidas por diferentes aceitadores de eletrões (nitrato vs sulfato). Assim, os extratos proteicos totais de células de D. desulfuricans crescidas em meios VMN com nitrato ou sulfato, foram analisados por eletroforese bidimensional (2DE), num gradiente de pH 4 - 7. Nas condições experimentais testadas, foram observados 604 e 519 spots de proteínas nos géis de células crescidas em meio contendo nitrato ou sulfato, respetivamente. Pela avaliação estatística foi possível observar aproximadamente 25 % de spots diferenciais. Os resultados obtidos sugerem que na presença de nitrato, a bactéria não só cresce mais rapidamente e com maior rendimento, como também produz uma maior quantidade de proteínas. Estes dados foram relacionados com a adaptação de D. desulfuricans ao substrato respiratório alternativo. Tal como esperado, nos ensaios das atividades enzimáticas das redutases do nitrito e do nitrato, foi possível observar maior atividade nos extratos das células crescidas em nitrato do que em sulfato. As actinobactérias marinhas do género Salinispora, têm vindo a ser exploradas como fontes de biofármacos naturais. Assim, na segunda parte deste trabalho, realizou-se um estudo preliminar, que pretendeu caracterizar as proteínas envolvidas na biossíntese destes compostos bioactivos em S. arenicola e de S. pacifica. Para tal, foi utilizada uma abordagem proteómica diferencial, baseada em 2DE. Surpreendentemente, os perfis proteicos das duas espécies mostraram-se bastante distintos, tendo-se identificado apenas 37 spots comuns entre os 650 observados no gel de S. arenicola e 510 no gel de S. pacifica.

Methanogens, sulphate and heavy metals: a complex system

Anaerobic digestion (AD) is a well-established technology used for the treatment of wastes and wastewaters with high organic content. During AD organic matter is converted stepwise to methane-containing biogasa renewable energy carrier. Methane production occurs in the last AD step and relies on methanogens, which are rather sensitive to some contaminants commonly found in wastewaters (e.g. heavy metals), or easily outcompeted by other groups of microorganisms (e.g. sulphate reducing bacteria, SRB). This review gives an overview of previous research and pilot-scale studies that shed some light on the effects of sulphate and heavy metals on methanogenesis. Despite the numerous studies on this subject, comparison is not always possible due to differences in the experimental conditions used and parameters explained. An overview of the possible benefits of methanogens and SRB co-habitation is also covered. Small amounts of sulphide produced by SRB can precipitate with metals, neutralising the negative effects of sulphide accumulation and free heavy metals on methanogenesis. Knowledge on how to untangle and balance sulphate reduction and methanogenesis is crucial to take advantage of the potential for the utilisation of biogenic sulphide as a metal detoxification agent with minimal loss in methane production in anaerobic digesters.

O efeito da insulina na placentação: estudos in vitro com células HTR8-SVneo

Dissertação de mestrado em Bioquímica Aplicada

Growth of sulfate reducing bacteria on the methanogenic inhibitor BES

The metabolism of methanogenic archaea is inhibited by 2-bromoethanesulfonate (BES). Methane production is blocked because BES is an analog of methyl-coenzyme M and competes with this key molecule in the last step of methanogenesis. For this reason, BES is commonly used in several studies to avoid growth of acetoclastic and hydrogenotrophic methanogens [1]. Despite its effectiveness as methanogenic inhibitor, BES was found to alter microbial communities’ structure, to inhibit the metabolism of non-methanogenic microorganisms and to stimulate homoacetogenic metabolism [2,3]. Even though sulfonates have been reported as electron acceptors for sulfate- and sulfite-reducing bacteria (SRB), only one study described the reduction of BES by complex microbial communities [4]. In this work, a sulfate-reducing bacterium belonging to Desulfovibrio genus (98 % identity at the 16S rRNA gene level with Desulfovibrio aminophilus) was isolated from anaerobic sludge after several successive transfers in anaerobic medium containing BES as sole substrate. Sulfate was not supplemented to the anaerobic growth medium. This microorganism was able to grow under the following conditions: on BES plus H2/CO2 in bicarbonate buffered medium; on BES without H2/CO2 in bicarbonate buffered medium; and on BES in phosphate buffered medium. The main products of BES utilization were sulfide and acetate, the former was produced by the reduction of sulfur from the sulfonate moiety of BES and the latter likely originated from the carbon backbone of the BES molecule. BES was found, in this study, to represent not only an alternative electron acceptor but also to serve as electron donor, and sole carbon and energy source, supporting growth of a Desulfovibrio sp. obtained in pure culture. This is the first study that reports growth of SRB with BES as electron donor and electron acceptor, showing that the methanogenic inhibitor is a substrate for anaerobic growth.

Effect of sulfate and iron (III) on LCFA degradation by a methanogenic community

[Excerpt] Under anaerobic conditions long chain fatty acids (LCFA) can be converted to methane by syntrophic bacteria and methanogenic archaea. LCFA degradation was also reported in the presence of alternative hydrogenotrophic partners, such as sulfate-reducing bacteria (SRB) and iron-reducing bacteria (IRB), which generally show higher affinity for H2 than methanogens and are more resistant to LCFA [1,2,3]. Their presence in a microbial culture degrading LCFA can be advantageous to reduce LCFA toxicity towards methanogens, although high concentrations of external electron acceptor (EEA) can lead to outcompetition of methanogens and cease methane production. In this work, we tested the effect of adding sub-stoichiometric concentrations of sulfate and iron(III) to methanogenic communities degrading LCFA. (...)

Sulphate-reducing bacteria associated with biocorrosion: a review

Biocorrosion means any process of corrosion in wich microorganisms are somehow involved. As far as the petroleum industry is concerned, the anaerobic type is the more important, with Sulphate-Reducing Bacteria (SRB) accouting for half of the described processes. SRB are obligate anaerobs that use sulphur, sulphate or other oxidized sulphur compounds as oxidizing agents when decomposing organic material. A typical product of SRB metabolism, hydrogen sulphide -H2S-, is extremely toxic. In the present work we review the literature on mechanisms underlying biocorrosive process in wich SRB are involved and summarize some of the ultrastructural and eletrochemical work developed using SRB obtained from water injection flow in wells located on PETROBRAS offshore marine plataforms, sampled directly in the field over metallic probes, or cultured under laboratory conditions. Biofilms develop when SRB adhere to inert surfaces. A high diversity of morphological types is found inside these biofilms. Their extracellular matrix is highly hydrated and mainly anionic, as shown by its avid reaction with cationic compounds like ruthenium red. We have noted that variations in iron contet lead to interesting changes in the ultrastructure of the bacterial cell coat and also in the rate of corrosion induced in metallic test cupons. Since routine methods to prevent and treat SRB contamination and biodeterioration involve the use of biocides that are toxic and always have some environmental impact, an accurate diagnosis of biocorrosion is always required prior to a treatment decision. We developed a method that detects and semi-quantifies the presence of living or dead SRB by using free silver potentials as an indicator of corrosive action by SRB-associated sulphides. We found a correlation between sulphide levels (determined either by spectrophotometry, or using a silver electrode -E(Ag)- that measured changes in free potentials induced by the presence of exogeneously added sulphide) and SRB concentration (enumerated by a culturing method). E (Ag) was characterized under a variety of conditions andwas found to be relatively immune to possible interference resulting from aeration of media or from the psence of iron corrosion products. The method offers a simple, rapid, and effective means of diagnosing biocorrosive processes prior to their control.

Studies on the biology of schistosomiasis with emphasis on the Senegal river basin

The construction of the Diama dam on the Senegal river, the Manantali dam on the Bafing river, Mali and the ensuing ecological changes have led to a massive outbreak of Schistosoma mansoni in Northern Senegal, associated with high intensity of infections, due to intense transmission, and the creation of new foci of S. haematobium. Data on the vectorial capacity of Biomphalaria pfeifferi from Ndombo, near Richard Toll, Senegal are presented with sympatric and allopatric (Cameroon) S. mansoni. Comparisons are made on infectivity, cercarial production, chronobiology of cercarial emergence and longevity of infected snails. Recent data on the intermediate host specificity of different isolates of S. haematobium from the Lower and Middle Valley of the Senegal river basin (SRB) demonstrate the existence of at least two strains of S. haematobium. The role of Bulinus truncatus in the transmission of S. haematobium in the Lower and Middle Valleys of the SRB is reviewed. Both S. haematobium and S. mansoni are transmitted in the same foci in some areas of the SRB.

Period-Luminosity-Colour distribution and classification of Galactic oxygen-rich LPVs I. Luminosity calibrations

The absolute K magnitudes and kinematic parameters of about 350 oxygen-rich Long-Period Variable stars are calibrated, by means of an up-to-date maximum-likelihood method, using HIPPARCOS parallaxes and proper motions together with radial velocities and, as additional data, periods and V-K colour indices. Four groups, differing by their kinematics and mean magnitudes, are found. For each of them, we also obtain the distributions of magnitude, period and de-reddened colour of the base population, as well as de-biased period-luminosity-colour relations and their two-dimensional projections. The SRa semiregulars do not seem to constitute a separate class of LPVs. The SRb appear to belong to two populations of different ages. In a PL diagram, they constitute two evolutionary sequences towards the Mira stage. The Miras of the disk appear to pulsate on a lower-order mode. The slopes of their de-biased PL and PC relations are found to be very different from the ones of the Oxygen Miras of the LMC. This suggests that a significant number of so-called Miras of the LMC are misclassified. This also suggests that the Miras of the LMC do not constitute a homogeneous group, but include a significant proportion of metal-deficient stars, suggesting a relatively smooth star formation history. As a consequence, one may not trivially transpose the LMC period-luminosity relation from one galaxy to the other.

Kaivosteollisuudessa syntyvän kipsin hyödyntäminen

Diplomityön tarkoituksena oli tutkia ja kehittää käyttökohde kaivosteollisuudessa syntyvälle märälle kipsisivuvirralle, joka sisältää metalliepäpuhtauksina alumiinia, rautaa ja mangaania ja jonka määrä on noin 1 000 000 t/a. Kirjallisuuden pohjalta tutkittiin aluksi mahdollisuutta hyödyntää kipsiaines asfaltti- ja sementtiteollisuuden raaka-aineena. Sementin joukkoon lisätään tavallisesti noin 5 p-% kipsiä, mutta harvinaisimpiin sementtilaatuihin sitä voidaan lisätä jopa 30 p-%. Tästä huolimatta vain pieni osa tutkimuksen kohteessa syntyvästä kipsisivuvirrasta voitaisiin hyödyntää tässä sovelluksessa. Lisäksi kipsisivuvirran sisältämät epäpuhtaudet täytyisi poistaa tai saattaa inaktiiviseen muotoon. Myöskään sen kosteuspitoisuus ei saisi olla suuri. Näin ollen tämän kipsisivuvirran hyödyntäminen asfaltti- ja sementtiteollisuuden lisäaineena ei ole mahdollista Seuraavaksi harkittiin kipsin kierrättämistä, jolloin yhtenä vaihtoehtona oli hajottaa kipsi termisesti rikkioksideiksi ja valmistaa niistä rikkihappoa. Taloudellisista syistä hajoamistuotteen on oltava rikkitrioksidia, josta voitaisiin veteen imeyttämällä valmistaa rikkihappoa. Kipsin hajottaminen termovaa´alla osoitti, että kipsi vaatii noin 1400 ºC:n lämpötilan ja haihtuvat komponentit ovat H2O, SO ja SO2, muttei SO3. Alempien oksidien muuttaminen rikkihapoksi vaatisi katalyyttisen hapetuksen, mikä olisi käytännössä liian kallista. Toisena vaihtoehtona kipsin kierrättämiseksi tutkittiin sen biologista pelkistämistä rikkivedyksi ja kalsiumhydroksidilietteeksi. Laboratoriossa Ca(OH)2-lietteestä valmistettiin hiilidioksidin avulla kalsiumkarbonaattia, jolloin päästiin 90 %:n kalsiumhydroksidin konversiossa. Lisäksi alumiinihydroksidi saatiin erotettua kipsilietteestä kokeellisesti hydrosyklonin avulla. Diplomityössä päädyttiin siihen, että sulfaatin biologinen pelkistäminen ja alumiinihydroksidin mekaaninen erotus jatkuvatoimisesti on varteenotettava vaihtoehto kipsisivuvirran hyödyntämiseksi.

Atividade de extratos de Arrabidaea chica (Humb. & Bonpl.) Verlot obtidos por processos biotecnológicos sobre a proliferação de fibroblastos e células tumorais humanas

Arrabidaea chica (H&B) Verlot is a plant popularly known as Pariri and this species is a known source of anthocyanins, flavonoids and tannins. This report describes an approach involving enzymatic treatment prior to extraction procedures to enhance A chica crude extract anticancer activity. Anticancer activity in human cancer cell lines in vitro using a 48 h SRB cell viability assay was performed to determine growth inhibition and cytotoxic properties. The final extraction yield without enzyme treatment was higher (24.28%) compared to the enzyme-treated material (19.03%), with an enhanced aglycones anthocyanin ratio as determined by HPLC- DAD and LC-MS with direct infusion.

Dynamic Analysis Model of Spherical Roller Bearings with Defects

Rolling element bearings are essential components of rotating machinery. The spherical roller bearing (SRB) is one variant seeing increasing use, because it is self-aligning and can support high loads. It is becoming increasingly important to understand how the SRB responds dynamically under a variety of conditions. This doctoral dissertation introduces a computationally efficient, three-degree-of-freedom, SRB model that was developed to predict the transient dynamic behaviors of a rotor-SRB system. In the model, bearing forces and deflections were calculated as a function of contact deformation and bearing geometry parameters according to nonlinear Hertzian contact theory. The results reveal how some of the more important parameters; such as diametral clearance, the number of rollers, and osculation number; influence ultimate bearing performance. Distributed defects, such as the waviness of the inner and outer ring, and localized defects, such as inner and outer ring defects, are taken into consideration in the proposed model. Simulation results were verified with results obtained by applying the formula for the spherical roller bearing radial deflection and the commercial bearing analysis software. Following model verification, a numerical simulation was carried out successfully for a full rotor-bearing system to demonstrate the application of this newly developed SRB model in a typical real world analysis. Accuracy of the model was verified by comparing measured to predicted behaviors for equivalent systems.

Análise descritiva quantitativa de edulcorantes em diferentes concentrações

Edulcorantes em solução, com a mesma equivalência de doçura, podem apresentar características sensoriais que os tornam diferentes entre si. O presente estudo teve como objetivo realizar Análise Descritiva Quantitativa de soluções de aspartame (APM), extrato de folhas de estévia (SrB) e mistura ciclamato/sacarina 2:1 (C/S) em diferentes níveis de doçura, ou seja, em equivalência de doçura a uma solução aquosa de sacarose a 3, 10, 20 e 30%. Onze provadores, pré-selecionados através de análise seqüencial, tendo como critério suas habilidades de discriminação, foram treinados após o levantamento da terminologia descritiva. Após o treinamento, os provadores foram selecionados através de seu poder de discriminação, reproducibilidade e concordância com a equipe no uso de escalas. Os termos descritivos dos edulcorantes, para todos os níveis de doçura, gerados através do método rede (Kelly's Repertory Grid Method) foram: doçura inicial, doçura residual, amargo inicial, amargo residual, residual de alcaçuz, corpo e acidez. Os resultados obtidos para cada nível de doçura foram analisados através de análise de variância, teste de Tukey e Análise de Componentes Principais. A análise descritiva foi efetiva em caracterizar o perfil sensorial dos edulcorantes em diferentes concentrações, evidenciando as mudanças no perfil com o aumento de suas concentrações.

Dificultades en la integración y cooperación de los cuerpos policiales de América Latina en la lucha contra el narcotráfico. Estudio de caso: AMERIPOL

La presente investigación analiza los principales retos que afectan la cooperación e integración de los cuerpos policiales latinoamericanos en la lucha contra el narcotráfico. Para realizar dicho análisis se toma como caso de estudio AMERIPOL, organización creada en el año 2007 y que actualmente constituye el único mecanismo de cooperación policial en el hemisferio americano. Se considera que la cooperación e integración de los cuerpos policiales en la lucha contra el narcotráfico en la región enfrentan dos retos principales: la disparidad de los marcos jurídicos entre los Estados y la descoordinación política. Estos retos suponen la existencia de factores que impiden una política de cooperación conjunta contra las drogas y por tanto, el desarrollo de actitudes aislacionistas que limitan la cooperación en contra de éste flagelo

La construcción de la política exterior de Rusia hacia Serbia: entre el pragmatismo y el Eurasianismo (2007-2015)

Esta monografía defenderá que la Rusia contemporánea cuenta con una ideología formada a partir de los elementos imperiales y religiosos previos a la Unión Soviética además de elementos políticos propios de la URSS como la colectividad y la oposición a occidente. La Rusia de Putin ha recobrado sus capacidades para defender sus intereses en el mundo frente a la injerencia de occidente, y en el caso de Serbia esto se hará evidente a través de la importancia de los lazos culturales e históricos reforzados por los vínculos económicos y estratégicos que representa la presencia de Rusia en Serbia. Los principales autores sobre los que se trabajará son Alexander Dugin y Alexander Panarin.

Dietary glycated protein modulates the colonic microbiota towards a more detrimental composition in ulcerative colitis patients and non-ulcerative colitis subjects

AIM: To investigate the effect of native, heated and glycated bovine serum albumin (BSA) on the ulcerative colitis (UC) and non-UC colonic microbiota in vitro. METHODS AND RESULTS: Continuous flow culture (CFC) models of the human colonic microbiota inoculated with faeces from UC and non-UC volunteers were maintained on BSA as growth substrate. Changes in bacterial populations and short-chain fatty acids were determined. UC and non-UC microbiota differed significantly in microbial populations, with elevated numbers of sulfate-reducing bacteria (SRB) and clostridia in the microbiota from UC patients. Compared with native BSA, glycated BSA modulated the gut microbiota of UC patients in vitro towards a more detrimental community structure with significant increases in putatively harmful bacteria (clostridia, bacteroides and SRB; P < 0.009) and decreases in dominant and putatively beneficial bacterial groups (eubacteria and bifidobacteria; P < 0.0004). The levels of beneficial short-chain fatty acids were significantly decreased by heated or glycated BSA, but were increased significantly by native BSA. CONCLUSION: The UC colonic microbiota maintained in CFC was significantly modified by glycated BSA. SIGNIFICANCE AND IMPACT OF THE STUDY: Results suggest that dietary glycated protein may impact upon the composition and activity of the colonic microbiota, an important environmental variable in UC.