A região de Caçapava do Sul, no Estado do Rio Grande do Sul, compreende, principalmente, a quadrícula de mesmo nome e abrange cerca de 11.000 Km². A referida região é caracterizada pela ocorrência de um batólito granítico (Granito Caçapava) com afloramento de 25 x 10 Km, orientado no sentido N-S, e sobre o qual assenta-se a cidade supracitada. O Granito Caçapava é circundado por um cinturão de metamorfitos pertencentes à Formação Vacacaí, Grupo Porongos (Pré-Cambriano Superior) (RIBEIRO et alii, 1966). Dentre os metamorfitos situa-se um corpo de rochas carbonatadas de aspecto lenticular, com contatos nítidos, interprenetrado po apófises graníticas e com afloramento de aproximadamente 17 Km². Foram estudadas amostras deste corpo de rochas por meio de microscopia, microssonda eletrônica, difratometria e espectometria de raios X e análise química por via úmida, com o objetivo de identificar e caracterizar os minerais carbonáticos e silicáticos. Por meio de técnicas de coloração, foi possível a distinção de dolomita (Do), calcita (Cc) e calcita ferrífera (Ccf). As análises modal e química indicaram predominância de carbonatos sobre silicatos, estando estes últimos relacionados mais às bandas da rocha. Os minerais silicáticos detectados foram: talco, tremolita, diopsídio, forsterita, clorita e flogopita. Em menores quantidades aparecem titanita, apatita e quartzo, sendo comum, também, a serpentina oriunda de alteração da forsterita. Lançados em mapa, os minerais índices mostraram a existência de um zoneamento metamórfico com rochas pertencentes à Fácies Piroxênio Hornfels, próximo ao contato, e de rochas da Fácies Albita-Epídoto Hornfels, em porções mais distantes, evidenciando assim, metamorfismo térmico. Com base em diagramas T-\'X GRAUS\'C \'O IND.2\' para o sistema CaO-MgO-Si \'O IND.2\'-\'CO IND.2\'-\'H IND. 2\'O, foi analisado o metamorfismo destas rochas e estimadas as temperaturas máximas da ordem de \'560 GRAUS\'C, compatíveis com as temperaturas magmáticas do Granito Caçapava. A conclusão final advinda com os estudos ora realizados, sobre essas rochas carbonáticas, aponta no sentido de se tratarem de MÁRMORES DOLOMÍTICOS IMPUROS DE CONTATO, gerados durante o Ciclo Brasiliano (680 a 500 m.a.). / Not available.

The modular polyketide synthase responsible for biosynthesis of the antibiotic mupirocin occupies 75 kb of Pseudomonas fluorescens NCIMB 10586, while a hybrid of PKS/NRPS is responsible for biosynthesis of the antibiotic thiomarinol located on a 97 kb plasmid pTML1 in Pseudoalteromonas spp SANK 73390. Biosynthesis of the acyl side chains in mupirocin and thiomarinol are thought to be either through esterification of the fully synthesised fatty acid (C\(_9\) or C\(_8\)) or through extension of the PK derived ester starter unit which is predicted to be carried out on MmpB and TmpB. mupU/O/V/C/F and macpE are proposed to be sufficient for the conversion of pseudomonic acid B to pseudomonic acid A. Mupirocin is regulated via quorum sensing, while regulation of thiomarinol was not identified. Production of thiomarinol was determined to occur after 8 hours of growth, while acidic conditions and use of acetone with ethyl acetate improved the extraction. TmlU, the thiomarinol amide ligase, did not complement a mupU mutant in mupirocin, and was found to block the biosynthesis of 9-hydroxynonanoic acid, causing truncation of 9-HN. This suggests that MupU, prevents MmpB from being an iterative PKS. KS-B2/ACP-B2 was shown to be involved in the removal of C8-OH from thiomarinol. Genetically manipulated mupU increased the production of mupirocin to 3 to 4 fold without abolishing PA-B production. Fused mupU-macpE complemented the NCIMB10586ΔmupUΔmacpE double mutant. However, insertion of this fusion into MmpB blocked the biosynthesis of mupirocin, while insertion after MmpA did not changed the pathway. Attempts to mobilise pTML1 revealed that a hybrid plasmid of RK2-R6K γ-ori was integrated into pTML1 but recovery of this cointegrate has not yet been recovered in E. coli.

Haemorrhage remains a leading cause of death in civilian and military environments. Recent research into emergency treatments for severe haemorrhaging injuries has resulted in production of a number of advanced haemostatic products. In certain scenarios, concomitant release of toxic materials may exacerbate trauma and ultimately reduce survival from such injuries. In particular, systemic absorption of the nerve agent VX (S-[2-(diisopropylamino)ethyl]-O-ethyl methylphosphonothioate) via wounds may rapidly cause muscle paralysis and death. This research explored the hypothesis that a haemostatic product could perform an additional function as a wound decontaminant. […] In summary, this project has successfully demonstrated that haemostatic products, based on an adsorptive mechanism of action, offer a highly effective means of countering the rapidly fatal effects of the chemical warfare agent VX present in wounded skin tissue. Further development of the concept of a haemostatic decontaminant has life-saving implications.

An analysis of over 34,000 free-text messages assigned by pharmacists to prescription orders over a 12-month period showed a sub-optimal exchange of information with the physician. Focus groups and observational research were conducted to provide a more in-depth understanding of the factors involved. The use of CPOE did not reduce opportunities for personal interaction. The capability to communicate electronically facilitated a non-interruptive workflow, beneficial for staff time and for limiting distractions. It also improved clinical documentation, which helped coordinate care of patients between members of the pharmacy team. However, the research identified several barriers to the effectiveness of communication via the CPOE system, including: the increased frequency of messages sent; poor display characteristics of the message; poor access to information to inform decision-making; one-way communication; and no assigned responsibility to respond. These factors need to be considered in the design of systems and supported by interprofessional training to optimise communication between the professionals.

There is a clear and well-established need for new antibiotics. The biosynthesis of the antibiotic mupirocin is an important model trans-AT polyketide synthase pathway. A key tailoring step is the removal of a hydroxyl from the intermediate pseudomonic acid B (PA-B), which generates active pseudomonic acid A (PA-A). This is proposed to occur on discrete proteins after release from enzyme MmpB. In this work, a systematic screen for genes required for this key step was developed, which implicated ten genes. On a multi-gene expression plasmid, these ten genes encoded all functions necessary for conversion. This re-identified seven genes from the existing proposed model. A novel requirement for MupM, MupN, and the DNA sequence but not protein function ofMupL was uncovered. This represents the first identification of all genes required for this surprisingly convoluted pathway. With the aim of re-engineering the location of these tailoring steps, domains from thiomarinol enzyme TmpB were inserted into MmpB. Low-level conversion to PA-A was achieved using thiomarinol tailoring enzymes. The TmpB KS0 domain was used to load the normally free-standing MacpE, and high-level conversion was achieved on the hybrid enzyme. This success opens the future path to further MmpB modifications to generate mupirocin derivatives.

Polyketide synthases (PKS) are enzyme complexes that synthesise many natural products of medicinal interest, notably a large number of antibiotics. The present work investigated the mupirocin biosynthesis system, comparing it with similar pathways such as thiomarinol and kalimantacin. The focus was on the structural modelling of the protein complexes involved in antibiotic synthesis, via molecular simulation and the analysis of structural and sequence data. Structural docking of acyl carrier proteins (ACP) cognate for an HMG-CoA synthase orthologue responsible for β-methylation (MupH) identified key residues involved in the recognitions specificity of the interacting partners, further supported by mutagenesis experiments, which thus allows prediction of β-methylation sites in PKS. Moreover, complementation and mutagenesis experiments performed on MupH homologs from kalimantacin and thiomarinol systems suggests specificity between the ACP:HCS proteins in the β-branching suggesting the possibility of engineering multiple specific β-branching modifications into the same pathway. Molecular dynamics simulations of ACPs from the mupirocin cluster revealed that the PKS ACPs form a cavity upon the attachment of the phosphopantetheine and acyl chains similar to what is seen in the fatty acid synthase ACPs and provide a better understanding of the structure function relationship in these small proteins. Molecular docking of the putative cognate substrate with the ketosynthase (KS) homo dimer of module 5 of the MmpA in the mupirocin pathway revealed a loop that may control specificity for the α-hydroxylated substrate and mutagenesis experiments support this proposition.

Protein kinases are major drug targets for many diseases. Among these are the STE20/SPS1-related proline/alanine-rich kinase (SPAK) and the oxidative- stress-responsive kinase 1 (OSR1), which are two related serine/threonine protein kinases. Both kinases are key components of the WNK-SPAK/OSR1 signalling pathway that has emerged as a key regulator of electrolyte homeostasis, body fluid and blood pressure. Various knock-in and knock-out SPAK and OSR1 mouse models exhibited reduced blood pressure. This highlighted SPAK and OSR1 kinases as promising targets in the treatment of hypertension. Encouraged by this, this project was initiated to discover specific WNK-signalling inhibitors by targeting SPAK and OSR1 kinases as potential novel antihypertensive agents. My work led to the identification of an allosteric pocket located in the highly conserved C-terminal domains of SPAK and OSR1, which influences their kinase activities. Using in silico screening, Rafoxanide, an anti-parasitic agent, was identified as a novel allosteric inhibitor of SPAK and OSR1. Additionally, high throughput screening led to the discovery of the clinically used agent, Verteporfin, as a novel and potent WNK-signalling inhibitor. Moreover, several fragment-binders to the C-terminal domain of OSR1 kinase were identified using NMRfragment based screening. In addition, the NMR backbone assignments of the C-terminal domain of OSR1 kinase were determined and used to map the previously unknown binding site of different OSR1 and SPAK inhibitors. Collectively, these findings have significantly advanced the field of SPAK and OSR1 kinase inhibition and provided key tools that will facilitate the future discovery of other SPAK and OSR1 kinase inhibitors.

Este estudo investiga a identidade profissional do professor particular de piano a partir da análise de sua trajetória docente, tendo como método de pesquisa a história oral. Através de entrevistas com 13 professores, de 62 a 79 anos de idade, atuantes na cidade de Porto Alegre, o trabalho analisa os vários processos de formação, as idéias, os valores e as concepções dos professores sobre a profissão, tomando como referencial teórico os estudos de NÓVOA (1995), HUBERMAN (1995) e SCHÕN (1983, 1995). Os resultados obtidos mostram que os professores particulares de piano que participaram desse trabalho possuem concepções claras sobre a profissão, assumem os limites e dificuldades encontrados no exercício dessa atividade profissional, e, sobretudo, estão atentos às mudanças que foram ocorrendo durante os anos de trajetória docente, indo ao encontro das necessidades do aluno em aprender música, avaliando e reajustando suas formas de ver e de proceder. Esse trabalho desmistifica a idéia de que o professor particular está descomprometido com seu papel como profissional.

Ambientes aquáticos são essenciais para quase todas as formas de vida na Terra. Suas comunidades microbianas são responsáveis por controlar processos biogeoquímicos importantes nestes ecossistemas. Nos últimos anos, ocorreu um considerável aumento no número de estudos relacionando fatores e processos ambientais e comunidades microbianas em ambientes aquáticos. Uma das principais razões para o crescimento destes estudos foram os avanços das tecnologias de sequenciamento de DNA, que permitiram o desenvolvimento de novas abordagens, proporcionando um momento único para o campo da microbiologia. A aplicação de sequenciamento de nova geração para analisar regiões do gene do RNA ribossomal 16S (rRNA 16S) tem sido amplamente utilizada para a compreensão da composição, estrutura e respostas ambientais do bacterioplâncton. Os rios são ambientes importantes, tanto em uma perspectiva ecológica quanto econômica. No entanto, a base de nosso conhecimento sobre a dinâmica microbiana aquática se baseia principalmente em estudos ambientais de água do mar e lagos. Nesta tese, foi realizado o sequenciamento em larga escala da região V4 do gene rRNA 16S em amostras coletadas ao longo do Rio dos Sinos. Este rio está localizado em um dos centros industriais brasileiros mais importantes e apresenta várias características ambientais contrastantes, além de um gradiente longitudinal de eutrofização, assim, tornando-se um local notável para o estudo da dinâmica do bacterioplâncton. Neste trabalho, foi testada a influência do espaço e do tempo como variáveis para o processo de estruturação das comunidades microbianas, bem como influências sazonais e longitudinais para suas dinâmicas ecológicas. Foram demonstradas evidências consistentes para a existência de um perfil bacteriano na nascente do rio e a sua persistência ao longo do curso de água analisado. Mudanças sazonais reforçam a importância da nascente do rio na manutenção do reservatório bacteriano que é dispersado ao longo de todo o rio. Além disso, também existe uma pequena influência do espaço para determinar a estrutura das comunidades microbianas onde, amostras separadas por 63 km de curso de água são quase tão semelhantes entre si quanto as amostras de um mesmo local, coletadas com uma diferença de 15 minutos. Portanto, a preservação da fonte do rio é importante não apenas por razões hidrológicas, mas também para manter a composição e a integridade microbiana do rio. / Aquatic environments are essential for almost all Earth living forms. Their microbial communities are responsible for driven the most important biogeochemical processes in these ecosystems. Only in the past few years, we began to understand about the factors and processes that orchestrate the microbial community in these environments. One reason for this delay was the lack of proper tools. With advances in new sequencing technologies, new approaches are providing an unique moment for the microbiology field. The application of next-generation sequencing for 16S rRNA analysis has been broadly used for understanding bacterioplankton composition, structure and environmental responses. Rivers are important environments, for both ecological and economic perspectives. However, the basis of our knowledge about aquatic microbial dynamics relies mostly in seawater and lakes environmental studies. We performed high-throughput analysis of 16S rRNA highly variable V4 region in samples from the Sinos River, which is located in one of most important Brazilian industrial centers. This region has several contrasts in its environmental characteristics, presenting a longitudinal gradient of eutrophication and making it a remarkable study site for observing the dynamics of bacterioplankton. In this work, we tested the influence of space and time as variables for the microbial community structuring process, as well as, seasonal and longitudinal influences for their ecological dynamics. We demonstrated consistent evidence for the existence of a bacterial seed-bank and its longitudinal persistence. Seasonal shifts reinforce the importance of the source of the river in maintaining the bacterial seed-bank that spreads throughout the river. Besides, our results also suggest a lower influence of the space to determine the microbial community structure, where, samples separated by 63 km of watercourse are almost as dissimilar as the samples from the same site, collected with a difference of fifteen minutes. Therefore, the preservation of the source of the river is important not only for hydrologic reasons but also to maintain the microbial composition and the ecological integrity of the river.

As bacias de captação Faxinal e Maestra são responsáveis pela maior parte do abastecimento público de água do Município de Caxias do Sul, RS, e se localizam em região recoberta pelas unidades ignimbríticas riodacíticas da Formação Serra Geral. O presente estudo relata a influência do uso do solo na composição geoquímica das águas e sedimentos finos observados nos principais tributários das bacias, tendo como objetivo aplicar a geoquímica multielementar e isotópica para determinar as concentrações e os diferentes tipos de fontes de contaminantes presentes nas bacias hidrográficas estudadas, desenvolvendo um diagnóstico capaz de auxiliar no planejamento e gestão destes recursos hídricos. Para tanto, foi necessário correlacionar a composição geoquímica dos sedimentos finos de fundo com a rocha-fonte e a composição dos principais fertilizantes utilizados nas áreas agrícolas. Adicionalmente, foram determinadas as razões isotópicas de Pb e Sr em sedimentos de suspensão, associando com as assinaturas isotópicas de fertilizantes e de sedimentos nas nascentes principais, relacionando com a influência dos insumos utilizados na agricultura e da estação de tratamento de esgoto (ETE) nas águas dos tributários, utilizando-se os isótopos de δ2H-H2O, δ18O-H2O, δ15N-NO3-, δ18O-NO3- e δ15N. As amostras de sedimentos, água e fertilizantes foram analisadas por difração de Raios-X, ICP-MS, espectrômetro de ionização térmica, espectrómetro de massa VG Sector 54, espectroscopia de absorção de laser (Tipo Cavity Ring-Down Spectroscopy) e espectrômetro de Massa por Razão Isotópica (IRMS). O primeiro manuscrito apresenta resultados obtidos nos sedimentos de fundo interpretados com suporte de estatística descritiva e multivariada (Análise de Componentes Principais e Análise Fatorial), relacionando com o fator de enriquecimento. Estes resultados demonstram que os sedimentos de corrente dos córregos da bacia de captação Faxinal expõem contaminação e efeito acumulativo por metais pesados provenientes da aplicação de fertilizantes e fungicidas, enquanto que o excesso de sódio nos sedimentos da bacia de captação Maestra são resultantes da descarga direta de esgoto doméstico no recurso hídrico. O segundo manuscrito expõe resultados de 87Sr/86Sr para sedimentos em suspensão (0,717324 – 0,723113) e para os fertilizantes (0,705 – 0,710) configurando duas populações. As razões isotópicas de chumbo para os sedimentos apresentaram valores de 208Pb/204Pb (38,42 – 38,97), 206Pb/204Pb (18,72 – 19,07), 207Pb/204Pb (15,60 – 15,75), 208Pb/206Pb (2,037 – 2,055) e 207Pb/206Pb (0,824 – 0,833). Os valores dos isótopos de estrôncio e chumbo demonstraram que os sedimentos em suspensão são compatíveis com origem geogênica, e apresentam influência de fungicidas utilizados nas áreas agrícolas. O terceiro manuscrito mostra resultados dos isótopos de N e O de nitrato nas águas demonstraram valores de δ15N entre 3,9 e 19,4, enquanto o δ18O entre 4,2 e 21,7. Estas assinaturas isotópicas estão representadas em diferentes compartimentos do gráfico característico δ15N-NO3- vs δ18O-NO3-. Os pontos monitorados Fx-01, Fx-02, Fx-03, Fx-04, Fx-05 e Fx-06 demonstram valores com influência e contribuição de fertilizantes. Por outro lado, o ponto Fx-07 sugere interferência por descarga direta de efluente doméstico no tributário e o ponto Fx-ETE apresenta assinatura característica de ambiente de denitrificação. Através da interpretação dos resultados obtidos nos três manuscritos reconhece-se que as águas das represas Maestra e Faxinal recebem contaminantes provenientes de diferentes fontes, que abrangem a aplicação de insumos agrícolas e as descargas de esgoto doméstico sem tratamento. / The Faxinal and Maestra catchment basin are responsible for most of the public water supply of the city of Caxias do Sul, RS, and are located in the region covered by riodacíticas ignimbríticas units of the Serra Geral Formation. This study describes the influence of land use in the geochemical composition of water and fine sediments observed in the main tributaries of the watershed, aiming to apply the multielement and isotopic geochemistry to determine the concentrations and the different types of sources of contaminants in the studied basins, developing a diagnosis to assist in the planning and management of water resources. Therefore, it was necessary to correlate the geochemical composition of botton fine sediment with the rock-source and composition of the main fertilizers used in the agricultural areas. Additionally, it was determined the isotopic ratios of Pb and Sr in the suspended sediment, associating with the isotopic signatures of fertilizers and sediment in the main springs, relating to the influence of the inputs used in agriculture and of the wastewater treatment plant (WWTP) in the tributaries of water, using the isotopes δ2H-H2O, δ18O-H2O, δ15N-NO3-, δ18O-NO3- and δ15N. The sediment samples, water and fertilizer were analyzed by X-ray diffraction, ICP-MS, Spectrometer thermal ionization, Mass spectrometer VG Sector 54, laser absorption spectroscopy (Type Cavity Ring-Down Spectroscopy), Isotopic ratio mass spectrometer (IRMS). The first manuscript presents results obtained in the bottom sediments interpreted with descriptive and multivariate statistical support (Principal Component Analysis and Factor Analysis), relating to the enrichment factor. These results demonstrate that the stream sediments of Faxinal watershed showed contamination and cumulative effect of heavy metals due to the application of fertilizers and fungicides, while the excess sodium in the sediments of Maestra watershed the result of direct discharge domestic sewage in the water resource. The second manuscript displays results of 87Sr / 86Sr for suspended sediment (0.717324 to 0.723113) and fertilizers (0.705 – 0.710) setting two populations. Lead isotopic ratios for sediments showed values of 208Pb/204Pb (38.42 – 38.97), 206Pb/204Pb (18.72 – 19.07), 207Pb/204Pb (15.60 – 15.75), 208Pb/206Pb (2.037 – 2.055) e 207Pb/206Pb (0.824 – 0.833). The values of strontium and lead isotopes showed that the suspended sediments are compatible with geogenic origin and present influence of fungicides used in agriculture areas. The third manuscript shows results of isotopes of C and O of nitrate in water showed δ15N values between 3.9 and 19.4, while δ18O between 4.2 and 21.7. These isotopic signatures are represented in different compartments of the characteristic chart δ15N-NO3- vs δ18O-NO3-. The points monitored Fx-01, Fx-02, Fx-03, Fx-04, Fx-05 and Fx-06 show values with influence and contribution of fertilizers. On the other hand, the Fx-07 point suggests interference by direct discharge of effluent household in the tributary and Fx-ETE point features characteristic signature of denitrification environment. Through the interpretation of the results obtained in the three manuscripts it is recognized that the waters of the Maestra and Faxinal dams receive contaminants from differents sources covering the application of agricultural inputs and domestic sewage discharges untreated.