Chlorophyllide a: Fact or Artifact - Resolution of the Chlorophyllide a Problem in the Routine Measurement of Planktonic Chlorophyll a

Chlorophyll a serves as the routine proxy for most estimates of phytoplankton biomass in limnology and oceanography. It is well known that enzymatic chlorophyllase activity breaks chlorophyll a into the degradation product chlorophyllide a (chlide a). This degradation could result in potentially large underestimates of the true chlorophyll a concentration when the analysis is made by modern chromatographic methods. The goal of this project was to determine proper protocol in the methodological elimination of artifactual chlorophyllide a formed during extraction of chlorophyll pigments through the application of a microwave-assisted solvent extraction technique. This study aimed to answer the question: Is chlorophyllide a a fact or artifact? Previous literature has suggested that chromatographically analyzed chlorophyllide a might be either an artifact of the extraction process or an in situ indicator of senescent, physiologically compromised phytoplankton due to environmental stressors. This study addresses this decades-old problem for chlorophyll analysis. The microwave technique described inhibits chlorophyllase enzymatic activity, preventing the artifactual production of chlorophyllide a. The heat of the microwave technique not only denatures the chlorophllyase enzyme activity, but also evaporates filter-retained water (known to promote enzyme activity in solvents such as acetone and methanol). This technique results in a sizeable increase in the yield of extracted chlorophyll a that was up to four times greater than the routine protocol of cold temperature solvent-soak technique using 90% acetone. Microwave-assisted extraction methodology consistently yielded higher concentrations of total chlorophyll a-like pigments than commonly employed solvent-soak technique used in long term sampling programs such as JGOFS, HOTS and BATS. The result of this study provides a method that 1) eliminates artifactual chlorophyllide a production and 2) increases the extraction yield of photosynthetic pigments in phytoplankton, including both chlorophylls and carotenoids. This new technique may have significant implications for estimates of oceanic primary production

Genome-Wide Transposon Screen of a Pseudomonas syringae mexB Mutant Reveals the Substrates of Efflux Transporters.

Bacteria express numerous efflux transporters that confer resistance to diverse toxicants present in their environment. Due to a high level of functional redundancy of these transporters, it is difficult to identify those that are of most importance in conferring resistance to specific compounds. The resistance-nodulation-division (RND) protein family is one such example of redundant transporters that are widespread among Gram-negative bacteria. Within this family, the MexAB-OprM protein complex is highly expressed and conserved among Pseudomonas species. We exposed barcoded transposon mutant libraries in isogenic wild-type and ΔmexB backgrounds in P. syringae B728a to diverse toxic compounds in vitro to identify mutants with increased susceptibility to these compounds. Mutants with mutations in genes encoding both known and novel redundant transporters but with partially overlapping substrate specificities were observed in a ΔmexB background. Psyr_0228, an uncharacterized member of the major facilitator superfamily of transporters, preferentially contributes to tolerance of acridine orange and acriflavine. Another transporter located in the inner membrane, Psyr_0541, contributes to tolerance of acriflavine and berberine. The presence of multiple redundant, genomically encoded efflux transporters appears to enable bacterial strains to tolerate a diversity of environmental toxins. This genome-wide screen performed in a hypersusceptible mutant strain revealed numerous transporters that would otherwise be dispensable under these conditions. Bacterial strains such as P. syringae that likely encounter diverse toxins in their environment, such as in association with many different plant species, probably benefit from possessing multiple redundant transporters that enable versatility with respect to toleration of novel toxicants.IMPORTANCE Bacteria use protein pumps to remove toxic compounds from the cell interior, enabling survival in diverse environments. These protein pumps can be highly redundant, making their targeted examination difficult. In this study, we exposed mutant populations of Pseudomonas syringae to diverse toxicants to identify pumps that contributed to survival in those conditions. In parallel, we examined pump redundancy by testing mutants of a population lacking the primary efflux transporter responsible for toxin tolerance. We identified partial substrate overlap for redundant transporters, as well as several pumps that appeared more substrate specific. For bacteria that are found in diverse environments, having multiple, partially redundant efflux pumps likely allows flexibility in habitat colonization

The Role of Bacillithiol in Gram-Positive Firmicutes

Significance: Since the discovery and structural characterization of bacillithiol (BSH), the biochemical functions of BSH-biosynthesis enzymes (BshA/B/C) and BSH-dependent detoxification enzymes (FosB, Bst, GlxA/B) have been explored in Bacillus and Staphylococcus species. It was shown that BSH plays an important role in detoxification of reactive oxygen and electrophilic species, alkylating agents, toxins, and antibiotics. Recent Advances: More recently, new functions of BSH were discovered in metal homeostasis (Zn buffering, Fe-sulfur cluster, and copper homeostasis) and virulence control in Staphylococcus aureus. Unexpectedly, strains of the S. aureus NCTC8325 lineage were identified as natural BSH-deficient mutants. Modern mass spectrometry-based approaches have revealed the global reach of protein S-bacillithiolation in Firmicutes as an important regulatory redox modification under hypochlorite stress. S-bacillithiolation of OhrR, MetE, and glyceraldehyde-3-phosphate dehydrogenase (Gap) functions, analogous to S-glutathionylation, as both a redox-regulatory device and in thiol protection under oxidative stress. Critical Issues: Although the functions of the bacilliredoxin (Brx) pathways in the reversal of S-bacillithiolations have been recently addressed, significantly more work is needed to establish the complete Brx reduction pathway, including the major enzyme(s), for reduction of oxidized BSH (BSSB) and the targets of Brx action in vivo. Future Directions: Despite the large number of identified S-bacillithiolated proteins, the physiological relevance of this redox modification was shown for only selected targets and should be a subject of future studies. In addition, many more BSH-dependent detoxification enzymes are evident from previous studies, although their roles and biochemical mechanisms require further study. This review of BSH research also pin-points these missing gaps for future research. Antioxid. Redox Signal. 28, 445–462

Os sistemas de restrições das liberdades à luz da constituição federal de 1988

Orientador: Vera Karam de ChueiriMonografia (graduação) - Universidade Federal do Paraná, Setor de Ciências Jurídicas, Curso de Graduação em DireitoEste trabalho se propõe a estudar a formação histórica do direito da liberdade e seu tratamento constitucional no Brasil, enfatizando o funcionamento e possíveis desvios dos sistemas de restrições das liberdades adotados na Constituição Federal de 1988. O primeiro ponto analisa a formação histórica do direito de liberdade. O segundo ponto se concentra no tratamento dado à liberdade nas Constituições brasileiras. O terceiro ponto se preocupa com os vários sentidos e formas atribuídos à liberdade, bem como com as disposições assecuratórias na ordem constitucional vigente. O ponto quarto apresenta uma noção geral de regras e princípios e de eficácia da norma jurídica, para então analisar os sistemas de restrições que a Constituição Federal adota e avaliar como se aperfeiçoaria a atuação do Poder Legislativo e o poder de polícia, a fim de se evitar o abuso de poder

Identification of altered function alleles that affect Bacillus subtilis PerR metal ion selectivity

Bacillus subtilis PerR is a Fur family repressor that senses hydrogen peroxide by metal-catalyzed oxidation. PerR contains a structural Zn(II) ion (Site 1) and a regulatory metal binding site (Site 2) that, upon association with either Mn(II) or Fe(II), allosterically activates DNA binding. In addition, a third less conserved metal binding site (Site 3) is present near the dimer interface in several crystal structures of homologous Fur family proteins. Here, we show that PerR proteins with substitutions of putative Site 3 residues (Y92A, E114A and H128A) are functional as repressors, but are unexpectedly compromised in their ability to sense H2O2. Consistently, these mutants utilize Mn(II) but not Fe(II) as a co-repressor in vivo. Metal titrations failed to identify a third binding site in PerR, and inspection of the PerR structure suggests that these residues instead constitute a hydrogen binding network that modulates the architecture, and consequently the metal selectivity, of Site 2. PerR H128A binds DNA with high affinity, but has a significantly reduced affinity for Fe(II), and to a lesser extent for Mn(II). The ability of PerR H128A to bind Fe(II) in vivo and to thereby respond efficiently to H2O2 was restored in a fur mutant strain with elevated cytosolic iron concentration

A utilização das mídias como nova forma de aprendizado e ensino para os professores de Educação Física do município de Palmeira

Orientador : Denise Eurich ColatussoArtigo (especialização) - Universidade Federal do Paraná, Setor de Educação Profissional e Tecnológica, Curso de Especialização em Mídias Integradas na Educação.Inclui referência

A percepção de gestores sobre o processo de inovação: um estudo de caso em uma cooperativa agroindustrialThe perception of managers about the innovation process: a case study in an agroindustrial cooperative

A inovação tem sido considerada um tema estratégico na administração, representando um maior desafio às empresas. No caso das empresas do setor cooperativista o tema inovação é muito relevante, tendo em vista a participação dessas empresas no desenvolvimento do Estado. Nesse sentido, este trabalho teve como objetivo analisar a gestão da inovação em uma cooperativa agroindustrial localizada no Oeste do Paraná, a partir da avaliação de fatores determinantes para o processo de inovação. Para o embasamento teórico foram abordados conceitos de inovação e gestão da inovação, ademais de fatores-chave à gestão da inovação. A metodologia consistiu em um estudo de caso, com abordagem quantitativa descritiva e aplicação de um questionário aos gestores da cooperativa. Os resultados deste estudo demonstram que a cooperativa estudada apresentou fatores em seu processo de inovação que podem estar relacionados com as características de uma empresa inovadora e empreendedora

QUEBRA DE DORMÊNCIA DE GEMAS EM PESSEGUEIRO COM PRODUTOS ALTERNATIVOS NO ALTO VALE DO ITAJAÍ

Reguladores de crescimento são importantes ferramentas a serviço da produção de frutas temperadas, quando bem delimitados os métodos, sua utilização favorece a eficiência produtiva. Por isso na busca de alternativas para o manejo produtivo da quebra de dormência em pessegueiro, conduziu-se o presente experimento lançando mão de extrato artesanal de alho e industrial de algas, urina bovina e Erger® frente ao manejo convencional com Cianamida Hidrogenada, cujos efeitos colaterais tem posto em questão a segurança na utilização deste produto. Os resultados demonstram o efeito análogo entre os compostos e a possibilidade de emprego de produtos alternativos em pessegueiros