Pyoverdine and proteases affect the response of pseudomonas aeruginosa to gallium in human serum

Gallium is an iron mimetic which has recently been repurposed as an antibacterial agent due to its capability to disrupt bacterial iron metabolism. In this study, the antibacterial activity of gallium nitrate [Ga(NO3)3] was investigated in complement-free human serum (HS) on 55 Pseudomonas aeruginosa clinical isolates from cystic fibrosis and non-cystic fibrosis patients. The susceptibility of P. aeruginosa to Ga(NO3)3 in HS was dependent on the bacterial ability to acquire iron from serum binding proteins (i.e., transferrin). The extent of serum protein degradation correlated well with P. aeruginosa growth in HS, while pyoverdine production did not. However, pyoverdine-deficient P. aeruginosa strains were unable to grow in HS and overcome iron restriction, albeit capable of releasing proteases. Predigestion of HS with proteinase K promoted the growth of all strains, irrespective of their ability to produce proteases and/or pyoverdine. The MICs of Ga(NO3)3 were higher in HS than in an iron-poor Casamino Acids medium, where proteolysis does not affect iron availability. Coherently, strains displaying high proteolytic activity were less susceptible to Ga(NO3)3 in HS. Our data support a model in which both pyoverdine and proteases affect the response of P. aeruginosa to Ga(NO3)3 in HS. The relatively high Ga(NO3)3 concentration required to inhibit the growth of highly proteolytic P. aeruginosa isolates in HS poses a limitation to the potential of Ga(NO3)3 in the treatment of P. aeruginosa bloodstream infections

Copper acquisition by the SenC protein regulates aerobic respiration in Pseudomonas aeruginosa PAO1

Aerobic respiration of Pseudomonas aeruginosa involves four terminal oxidases belonging to the heme-copper family (that is, three cytochrome c oxidases and one quinol oxidase) plus one copper-independent, cyanide-insensitive quinol oxidase (CIO). The PA0114 gene encoding an SCO1/SenC-type protein, which is known to be important for copper delivery to cytochrome c in yeast, Rhodobacter spp. and Agrobacterium tumefaciens, was found to be important for copper acquisition and aerobic respiration in P. aeruginosa. A PA0114 (senC) mutant grew poorly in low-copper media and had low cytochrome cbb3-type oxidase activity, but expressed CIO at increased levels, by comparison with the wild-type PAO1. Addition of copper reversed these phenotypes, suggesting that periplasmic copper capture by the SenC protein helps P. aeruginosa to adapt to copper deprivatio

3D printed clotrimazole intravaginal ring for the treatment of recurrent vaginal candidiasis

Vulvovaginal candidiasis is a vaginal infection caused by the fungal pathogen Candida albicans that, most commonly, affects women of reproductive age. Its first-line treatment consists in topical applications of conventional drug formulations (e.g., creams, gels, tablets) containing imidazole drugs. The treatment involves single or multiple daily applications and, in the case of recurrences, daily administration of oral antifungal drugs for up to one month. Intravaginal rings are flexible, biocompatible medical devices that, compared to conventional drug formulations, offer the possibility of a controlled vaginal drug delivery over a determined period with a single application, thus increasing patient compliance. Among innovative manufacturing techniques, in recent years, fused deposition modeling 3D printing has emerged in the pharmaceutical field to produce different therapeutics combining drugs and polymers. This technique allows to print objects layer by layer with many different thermoplastic materials after a computer-assisted design. Thermoplastic polyurethanes are flexible and biocompatible materials that can be efficiently employed for the manufacturing of drug release systems, already utilized to prepare vaginal devices. In this work, we produced a clotrimazole-loaded intravaginal ring by fused deposition modeling 3D printing combining the drug with thermoplastic polyurethane using hot melt extrusion. The rings were computer-designed and then printed with two different drug concentrations (i.e., 2% and 10% w/w). The intravaginal rings were first tested in an agar-diffusion test to evaluate their effectiveness against C. albicans; and the 10% loaded ring was selected for further studies. Drug release was evaluated in two different media (i.e., 50% ethanol and vaginal fluid simulant) showing a sustained release over a period of seven days. Next, in vitro time-kill analysis against C. albicans in simulated vaginal fluid was performed and displayed a complete growth inhibition after 5 days, compared to the control. These results suggest a potential application of these 3D printed intravaginal rings for the treatment of vulvovaginal candidiasis and for the long-time treatment of recurrences

Padrões de composição e diversidade funcionais de florestas do pampa no sul do Brasil

Ecólogos de comunidades têm como um dos principais objetivos identificar padrões de abundância e distribuição de espécies ao longo das regiões e ambientes, e inferir sobre os processos norteadores desses padrões. Para tanto, duas linhas teóricas complementares são principalmente utilizadas: limitação de similaridade e filtros ambientais. Enquanto a primeira prediz que as espécies dentro de uma comunidade devem ser distintas nas suas características como resultado da competição (divergência), a segunda espera uma alta similaridade entre espécies coexistentes (convergência), uma vez que elas são filtradas sob certas condições ambientais. Estudos sobre diversidade funcional podem nos dar uma ideia da proporção de divergência e convergência na estruturação das comunidades, enquanto a composição funcional pode indicar a direção das respostas funcionais aos filtros ambientais. Ambientes campestres predominantemente cobrem o bioma Pampa, no sul do Brasil, e nesta região as florestas são restritas a zonas ribeirinhas e a pequenas encostas. As inundações são uma pressão comum em matas ribeirinhas, mas dadas as condições da região elas também são vulneráveis à dessecação por evapotranspiração e ventos. Em contraste, as florestas de encosta são mais sombreadas e úmidas. No presente trabalho, temos como objetivo principal caracterizar a diversidade e composição funcional destes ambientes florestais. Esperamos encontrar padrões de convergência para os atributos de florestas ciliares, devido às condições ambientais mais severas. Estabelecemos 18 unidades amostrais de 250 x 10 m em três regiões do Pampa, nove para cada tipo de floresta. Todas as árvores com pelo menos 5 cm de DAP (diâmetro à altura do peito) foram medidas. As regiões representam um gradiente de precipitação e temperatura. Seis atributos funcionais das espécies arbóreas foram avaliados: área foliar, área foliar específica (SLA), conteúdo de matéria seca foliar (LDMC), altura máxima, síndrome de dispersão e tamanho do fruto. Índices de diversidade funcional e seus tamanhos de efeito padronizados (SES - standardized effect size) dada a riqueza local de espécies foram estimados para cada região e tipo florestal. A composição funcional foi analisada através de média dos atributos funcionais ponderada pelas abundâncias nas comunidades (CWM), com posterior análise de variância para verificar diferenças entre as florestas e regiões. Os valores de CWM de área foliar e proporção de zoocoria foram menores nas florestas ribeirinhas de todas as regiões, enquanto o tamanho do fruto, SLA e LDMC só foram influenciados pelo fator região, sendo a SLA menor na região de menor precipitação e temperatura médias anuais. Altura máxima média das comunidades diferiu tanto conforme o tipo de floresta (maior na encosta) quanto à região (maior na região com condições climáticas mais favoráveis). Os valores do SES para riqueza funcional e dispersão funcional não demostraram padrões de divergência ou convergência significativos. Apenas a riqueza funcional para altura e tamanho de fruto foi distinta de acordo com o ambiente e a região das florestas avaliadas. Os resultados deste trabalho, ao contrário do esperado, demonstram não haver padrões de divergência e/ou convergência funcional para as florestas do Pampa, em termos do ambiente em que ocorrem (ciliar ou encosta) ou mesmo região. Contudo, a composição funcional variou tanto em relação ao ambiente quanto à região. Isso pode indicar uma seleção de espécies com distintas estratégias adaptativas de acordo com as diferentes influências ambientais em escala local e regional.Community ecologists aim at identifying patterns of species abundance and distribution along regions and environments, inferring on their main driver processes. Two complementary approaches rule community assembly: limiting similarity and environmental filters. While the first predicts that species within a community should be distinct in traits to avoid competition (divergence), the second expects a high similarity between co-occurring species (convergence), since they are filtered out to occur under certain environmental conditions. Functional diversity can give us an idea of the proportion of divergence and convergence in the structure of communities, while the functional composition might indicate the direction of trait responses to the environmental filters. Grasslands predominantly cover the southern Brazilian Pampa, and the forests are restricted to riparian zones and hillslopes. Flooding is a common pressure on riparian forests, but given the environmental conditions of the region they are also vulnerable to desiccation by evapotranspiration and winds. In contrast, slope forests are more shaded and humid. We aimed at characterizing functional diversity and composition of these forest environments. We expected trait convergence for riparian forests because of the harsher environmental conditions. We established 18 sampling units of 250 x 10 m in three regions of the Pampa, nine for each forest type. All trees with at least 5cm DBH were measured. The regions represent a gradient in precipitation and temperature. Six functional traits of the tree species were assessed: leaf area, specific leaf area (SLA), leaf dry matter content (LDMC), plant height, dispersal mode, and fruit size. Functional diversity indices and their standardized effect size (SES), considering the local species richness, were estimated for each region and forest. Functional composition was analyzed through community-weighted mean of traits (CWM), with further analysis of variance to verify differences between forests and regions. CWM of leaf area and zoochory were lower in riparian forests of all regions, while fruit size, SLA and LDMC were only influenced by the region. SLA was lower in the region with lower annual mean precipitation and temperature. Maximum height shows response for the type of forest (higher CWM in hillslope forests) and region (higher in the region under more favorable climatic conditions). SES of functional richness and dispersion did not show a recurrent pattern of significant divergence or convergence related to the factors. The results of SES, contrary to our expectation, are not indicating any patterns of trait divergence and/or convergence for the forests. Only the functional richness of height and fruit size differed according to the habitat and the region of the studied forests. However, both forest environments and regions differed in terms of functional composition, indicating the assembling of species with distinct adaptive strategies in relation to different environment influences for local and regional scale

Recuperação de poli (3-hidroxibutirato) produzido por Priestia megaterium utilizando cloreto de 1-hexadecilimidazólio

Polihidroxialcanoatos (PHAs) são poliésteres biodegradáveis que apresentam potencial para serem aplicados em substituição a polímeros de origem petroquímica por possuírem características térmicas e mecânicas semelhantes a estes. O poli(3-hidroxibutirato) [P(3HB)] é um dos PHAs mais estudados por sua versatilidade de produção e aplicação. Esse polímero é produzido por algumas espécies bacterianas e, então, pode ser recuperado do interstício celular por métodos mecânicos, químicos ou biológicos. Destaca-se, dentre estes, a utilização de solventes orgânicos por ser o mais utilizado. O principal destes solventes é o clorofórmio, que é tóxico e nocivo ao meio ambiente; sendo, portanto, de grande importância a busca de alternativas mais sustentáveis e menos prejudiciais. Neste contexto, líquidos iônicos (LIs), que são sais que se apresentam líquidos a temperaturas de 100 ºC ou menores e apresentam características que possibilitam a solubilização de uma série de compostos, surgem como uma alternativa para a obtenção deste biopolímero do interior das células. O objetivo do presente trabalho foi de avaliar o desempenho do LI cloreto de 1-hexadecilimidazólio na recuperação de P(3HB) do interior de células da espécie Gram-positiva Priestia megaterium. Para tanto, primeiramente, realizou-se a validação de um método para a quantificação do biopolímero, baseado na digestão do P(3HB) com ácido sulfúrico concentrado e análise do ácido crotônico gerado por HPLC. Em sequência, realizou-se um teste para verificar uma possível interação entre as soluções de LI de concentrações 1 mg/mL e 100 mg/mL e nas temperaturas de 30, 60 e 90 °C com amostras de biopolímero puro adquirido comercialmente. Enfim, um teste de contato entre 0,1 g de células de P. megaterium e 1 mL de soluções de LI com concentrações de 1, 10 e 100 mg/mL foi realizado nas temperaturas de 30 °C e 60 °C a fim de avaliar a capacidade de recuperação do biopolímero do interior das células bacterianas, bem como a pureza do produto obtido desta maneira. A partir dos experimentos, foi possível constatar uma boa resposta do método de quantificação do P(3HB) a partir do HPLC, sendo os resultados obtidos próximos àqueles alcançados a partir de outras metodologias. Além disso, os resultados de contato entre as soluções de LI e o biopolímero puro apresentaram a possibilidade de coprecipitação do LI juntamente ao P(3HB), se apresentando como uma fonte de impurezas para o biopolímero recuperado. Para os testes de contato das soluções de LI com as células bacterianas obteve-se o melhor desempenho para a solução menos concentrada na menor temperatura, na qual a pureza média entre as amostras obtida foi de 56,4 % e a recuperação de 91,4 %.Polyhydroxyalkanoates (PHAs) are biodegradable polyesters that have the potential to be applied to replace polymers of petrochemical origin because they have thermal and mechanical characteristics similar to these. Poly(3-hydroxybutyrate) [P(3HB)] is one of the most studied PHAs due to its production and application versatility. This polymer is produced by some bacterial species and then can be recovered from the cell interstitium by mechanical, chemical or biological methods. Among these, the use of organic solvents stands out because it is the most used. Chief among these solvents is chloroform, which is toxic and harmful to the environment, therefore the search for more sustainable and less harmful alternatives is of great importance. In this context, ionic liquids (ILs), which are salts that are liquid at temperatures of 100 ºC or lower and have characteristics that enable the solubilization of a series of compounds, emerge as an alternative for obtaining this biopolymer from inside cells. The objective of the present work was to evaluate the performance of LI 1-hexadecylimidazolium chloride in the recovery of poly(3-hydroxybutyrate) from the interior of cells of the Gram-positive species Priestia megaterium. To do so, first, the validation of the method for the quantification of the biopolymer was carried out, based on the digestion of P(3HB) with concentrated sulfuric acid and analysis of the crotonic acid generated by HPLC. In sequence, a test was carried out to verify a possible interaction between the IL solutions of concentrations 1 mg/mL and 100 mg/mL and at temperatures of 30, 60 and 90 °C with samples of pure biopolymer purchased commercially. Finally, a contact test between 0.1 g of P. megaterium cells and 1 mL of IL solutions with concentrations of 1, 10 and 100 mg/mL was performed at temperatures of 30 °C and 60 °C in order to evaluate the recovery capacity of the biopolymer inside the bacterial cells, as well as the purity of the product obtained in this way. From the experiments, it was possible to verify a good response of the method of quantification of P(3HB) from the HPLC, being the results obtained close to those reached from other methodologies. In addition, the contact results between the IL solutions and the pure biopolymer showed the possibility of IL co-precipitation together with P(3HB), presenting itself as a source of impurities for the recovered biopolymer. For contact tests of IL solutions with bacterial cells, the best performance was obtained for the less concentrated solution at the lowest temperature, in which the average purity between the samples obtained was 56.4% and the recovery was 91.4 %

Estudo do tensofissuramento em geomembranas de polietileno de alta densidade utilizadas em aterros de resíduos

As geomembranas de polietileno de alta densidade (PEAD) são amplamente utilizadas em obras na área de meio ambiente como barreira impermeabilizante para evitar a contaminação das águas subterrâneas, pelas suas características de alta resistência química, flexibilidade e baixa permeabilidade a líquido e gases. Baseando-se no fato de que o aterro sanitário é recomendado atualmente como a melhor alternativa para o destino final das milhares de toneladas de resíduos sólidos gerados diariamente nas cidades e que, geomembranas de PEAD são utilizadas como reforço de impermeabilização nestas estruturas, o presente trabalho tem como objetivo avaliar as perdas de propriedade causadas pelas condições as quais a geomembrana encontra-se submetida quando instalada na célula de um aterro sanitário. Para tanto foram retiradas amostras da geomembrana impermeabilizante de um aterro sanitário que se encontrava sob resíduos há aproximadamente 1 ano. Estas amostras, juntamente com amostra de uma geomembrana nova, disponibilizada no próprio aterro, foram submetidas a dois tipos de ensaios: o primeiro ensaio, tensofissuramento ambiental de corpo de prova dobrado (Bent Strip Environmental Stress Cracking). Neste ensaio algumas amostras foram banhadas em solução tensoativa, conforme preconiza a norma, enquanto outras foram ensaiadas banhadas em chorume, numa adaptação à metodologia normalizada. No segundo ensaio, corpos de prova obtidos das amostras das geomantas foram submetidos a processo de envelhecimento parte em banho com solução tensoativa e parte com chorume. Após ambos os ensaios, as amostras e corpos de prova foram caracterizadas por ensaios térmicos, físicos, mecânicos, reológicos e morfológicos. No ensaio de tensofissuramento de corpo de prova dobrado não foi possível identificar a propagação de trinca ou fratura nas amostras porém foi possível identificar por MEV o início de microfissuras. Com base nos resultados reológicos e de tração modificado (gravatas) foi possível comprovar o processo de degradação química do polímero que ocorre pelas condições em que a geomembrana é submetida quando instalada em uma célula de aterro sanitário, notadamente sua porção localizada no talude lateral do aterro.High-density polyethylene geomembranes (HDPE) are currently widely used in environmental work as a waterproofing barrier to prevent contamination of groundwater, due to its characteristics of high chemical resistance, flexibility and low permeability to liquid and gases. Based on the fact that the landfill is currently recommended as the best alternative for the final destination of the thousands of tons of solid waste generated daily in our cities and that HDPE geomembranes are widely used as waterproofing reinforcement in these structures, the present work has as objective to evaluate properties losses caused by the conditions to which the geomembrane is submitted when installed in a cell of a landfill. For that purpose, exhumed samples from the cell of a sanitary landfill that had been under residues for approximately 1 year were obtained. These samples, together with a sample of a new geomembrane, made available in the own landfill, were submitted to two types of tests: The first one was bent strip environmental stress cracking test. In this test, some samples were bathed in a surfactant solution, according to the norm, while others were bathed in landfill leachate in an adaptation to the standard methodology. In the second test, test bodies obtained from the geosynthetics samples were submitted to aging process, part in a bath with surfactant solution and part with landfill leachate. After both tests, the test bodies, obtained from the geosynthetics samples, were characterized by thermal, physical, mechanical, rheological and morphological tests. In the bent strip environmental stress cracking test there was no possible to identify the propagation of crack or fracture in the samples; however it was possible to identify with scanning electron microscopy the beginning of microcracks. Based on the rheological and modified traction (ties) results it was possible to prove the chemical degradation process of the polymer that occurs due to the conditions in which the geomembrane is submitted when installed in a landfill cell, especially in its portion installed in the side embankment of the landfill

Draft Genome Sequence of the Carboxydotrophic Alphaproteobacterium Aminobacter carboxidus Type Strain DSM 1086

Aminobacter carboxidus is a soil Gram-negative alphaproteobacterium belonging to the physiological group of carboxydobacteria which aerobically oxidize CO to CO2. Here, we report the draft genome sequence of the A. carboxidus DSM 1086 type strain and the identification of both form I and form II CO dehydrogenase systems in this strain

Draft Genome Sequence of the Carboxydotrophic Alphaproteobacterium Aminobacter carboxidus Type Strain DSM 1086

Aminobacter carboxidus is a soil Gram-negative alphaproteobacterium belonging to the physiological group of carboxydobacteria which aerobically oxidize CO to CO2 Here, we report the draft genome sequence of the A. carboxidus DSM 1086 type strain and the identification of both form I and form II CO dehydrogenase systems in this strain

Draft genome sequence and secondary metabolite biosynthetic potential of the lysobacter niastensis type strain DSM 18481

Lysobacter niastensis belongs to a group of bacterial predators that produce a number of bioactive small molecules endowed with lytic properties toward other microorganisms. Here, we report the draft genome sequence of the type strain DSM 18481 and the identification of gene clusters implicated in the biosynthesis of secondary metabolites