Study of DNA damage with a new system for irradiation of samples in a nuclear reactor

In this paper, we report results of a quantitative analysis of the effects of neutrons on DNA, and, specifically, the production of simple and double breaks of plasmid DNA in aqueous solutions with different concentrations of free-radical scavengers. The radiation damage to DNA was evaluated by electrophoresis through agarose gels. The neutron and gamma doses were measured separately with thermoluminescent detectors. In this work, we have also demonstrated usefulness of a new system for positioning and removing samples in channel BH#3 of the IEA-R1 reactor at the Instituto de Pesquisas Energeticas e Nucleares (Brazil) without necessity of interrupting the reactor operation. (C) 2010 Elsevier Ltd. All rights reserved.

The Sphene-Centered Ocellar Texture: An Effect of Grain-Supported Flow and Melt Migration in a Hyperdense Magma Mush

The sphene-centered ocellar texture consists of leucocratic ocelli with sphene (titanite) crystals at the center, enclosed in a biotite-rich matrix. This texture has been recognized worldwide in hybrid intermediate rocks. On the basis of structural, petrological, and geochronological data from selected outcrops of the Variscan Ribadelago pluton (NW Iberian Massif), we propose that the ocelli were formed by migration and accumulation of a residual melt through a plagioclase- and biotite-dominated crystalline framework. At the late stage of crystallization, the magma acted as a hyperdense suspension and reacted to the pressure gradient caused by the regional stress field, entering the domain of grain-supported flow. Microstructures reveal that aligned crystal domains arose in the crystal framework from the shearing and compaction of the crystal mush and behaved as magmatic microshears. Relative displacement of adjacent crystal clusters along these microshears corresponded to the onset of Reynolds dilatancy that generated an expansion of the crystal mush, involving melt migration and pore aperture. The mineralogy of the ocelli, dominated by andesine and sphene, represents the composition of the migrating melt. The chemistry of this late, Ti-rich melt stems from the incongruent melting of biotite. Magmatic sphene from the ocelli yields a U-Pb age of 317 +/- 1 Ma, which represents the final crystallization of the hybridized magmatic system. Moreover, this texture offers an opportunity to better understand the rheological behavior of highly crystallized magmas.

Versatile microanalytical system with porous polypropylene capillary membrane for calibration gas generation and trace gaseous pollutants sampling applied to the analysis of formaldehyde, formic acid, acetic acid and ammonia in outdoor air

The analytical determination of atmospheric pollutants still presents challenges due to the low-level concentrations (frequently in the mu g m(-3) range) and their variations with sampling site and time In this work a capillary membrane diffusion scrubber (CMDS) was scaled down to match with capillary electrophoresis (CE) a quick separation technique that requires nothing more than some nanoliters of sample and when combined with capacitively coupled contactless conductometric detection (C(4)D) is particularly favorable for ionic species that do not absorb in the UV-vis region like the target analytes formaldehyde formic acid acetic acid and ammonium The CMDS was coaxially assembled inside a PTFE tube and fed with acceptor phase (deionized water for species with a high Henry s constant such as formaldehyde and carboxylic acids or acidic solution for ammonia sampling with equilibrium displacement to the non-volatile ammonium ion) at a low flow rate (8 3 nLs(-1)) while the sample was aspirated through the annular gap of the concentric tubes at 25 mLs(-1) A second unit in all similar to the CMDS was operated as a capillary membrane diffusion emitter (CMDE) generating a gas flow with know concentrations of ammonia for the evaluation of the CMDS The fluids of the system were driven with inexpensive aquarium air pumps and the collected samples were stored in vials cooled by a Peltier element Complete protocols were developed for the analysis in air of NH(3) CH(3)COOH HCOOH and with a derivatization setup CH(2)O by associating the CMDS collection with the determination by CE-C(4)D The ammonia concentrations obtained by electrophoresis were checked against the reference spectrophotometric method based on Berthelot s reaction Sensitivity enhancements of this reference method were achieved by using a modified Berthelot reaction solenoid micro-pumps for liquid propulsion and a long optical path cell based on a liquid core waveguide (LCW) All techniques and methods of this work are in line with the green analytical chemistry trends (C) 2010 Elsevier B V All rights reserved

Fluorescent amplified fragment length polymorphism genotyping of human and animal Staphylococcus aureus isolates from dairy farms with manual milking

Fluorescence amplified fragment length polymorphism (fAFLP) was used to assess the genetic relatedness of 40 Staphylococcus aureus strains isolated from human and animal skin samples in seven dairy farms with manual milking. S. aureus was isolated from 11 out of 30 (36%) human skin samples and from 29 out of 100 (29%) teat skin samples from apparently healthy cows. Genomic DNA from each isolate was double-digested with EcoRI and MseI and complementary oligonucleotide adaptors were ligated to the restriction fragments. Pre-selective and selective, amplification reactions were performed, the amplified fragments were separated by electrophoresis in an ABI377 sequencer and analysed using GeneScan 3.1 and Genotyper 2.5. Three single isolates (a-c), a predominant cluster with 35 isolates (d) and another cluster with two isolates (e) were identified. Both clusters d and e included human and animal isolates genetically related, because the profiles had 90-100% homology. Since no cluster was comprised uniquely of human or animal isolates and given the close genetic relatedness among human and animal samples in the farms, the present findings support the. hypothesis that dairy workers can spread S. aureus through manual milking. (C) 2005 Elsevier B.V. All rights reserved.

Genetic differentiation between sympatric populations of Bacillus cereus and Bacillus thuringiensis

Little is known about genetic exchanges in natural populations of bacteria of the spore-forming Bacillus cereus group, because no population genetics studies have been performed with local sympatric populations. We isolated strains of Bacillus thuringiensis and B. cereus from small samples of soil collected at the same time from two separate geographical sites, one within the forest and the other at the edge of the forest. A total of 100 B. cercus and 98 B. thuringiensis strains were isolated and characterized by electrophoresis to determine allelic composition at nine enzymatic loci. We observed genetic differentiation between populations of B. cereus and B. thuringiensis. Populations of a given Bacillus species-B. thuringiensis or B. cereus-were genetically more similar to each other than to populations of the other Bacillus species. Hemolytic activity provided further evidence of this genetic divergence, which remained evident even if putative clones were removed from the data set. Our results suggest that the rate of gene flow was higher between strains of the same species, but that exchanges between B. cereus and B. thuringiensis were nonetheless possible. Linkage disequilibrium analysis revealed sufficient recombination for B. cereus populations to be considered panmictic units. In B. thuringiensis, the balance between clonal proliferation and recombination seemed to depend on location. Overall, our data indicate that it is not important for risk assessment purposes to determine whether B. cereus and B. thuringiensis belong to a single or two species. Assessment of the biosafety of pest control based on B. thuringiensis requires evaluation of the extent of genetic exchange between strains in realistic natural conditions.

Sexagem de espermatozoides bovinos por centrifugação em gradiente descontínuo de densidade de Percoll

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ribotyping and virulence markers of Yersinia pseudotuberculosis strains isolated from animals in Brazil

Ribotyping and virulence markers has been used to investigate 68 Yersinia pseudotuberculosis strains of serogroups O:1a and O:3. The strains were isolated from clinical material obtained from healthy and sick animals in the Southern region of Brazil. Ribotypes were identified by double digestion of extracted DNA with the restriction endonucleases SmaI and PstI, separation by electrophoresis and hybridization with a digoxigenin-labeled cDNA probe. The presence of the chromosomal virulence marker genes inv, irp1, irp2, psn, ybtE, ybtP-ybtQ, and ybtX-ybtS, of the IS100 insertion sequence, and of the plasmid gene lcrF was detected by polymerase chain reaction. The strains were grouped into four distinct ribotypes, all of them comprising several strains. Ribotypes 1 and 4 presented distinct profiles, with 57.3% genetic similarity, ribotypes 2 and 3 presented 52.5% genetic similarity, and genetic similarity was 45% between these two groups (1/4 and 2/3). All strains possessed the inv, irp1, and irp2 genes. Additionally, strains of serogroup O:1a carried psn, ybtE, ybtP-ybtQ, ybtX-ybtS, and IS100. As expected lcrF was only detected in strains harboring the virulence plasmid. These data demonstrate the presence of Y. pseudotuberculosis strains harboring genotypic virulence markers in the livestock from Southern Brazil and that the dissemination of these bacteria may occur between herds.

Heart failure alters matrix metalloproteinase gene expression and activity in rat skeletal muscle

Heart failure is associated with a skeletal muscle myopathy with cellular and extracellular alterations. The hypothesis of this investigation is that extracellular changes may be associated with enhanced mRNA expression and activity of matrix metalloproteinases (MMP). We examined MMP mRNA expression and MMP activity in Soleus (SOL), extensor digitorum longus (EDL), and diaphragm (DIA) muscles of young Wistar rat with monocrotaline-induced heart failure. Rats injected with saline served as age-matched controls. MMP2 and MMP9 mRNA contents were determined by RT-PCR and MMP activity by electrophoresis in gelatin-containing polyacrylamide gels in the presence of SDS under non-reducing conditions. Heart failure increased MMP9 mRNA expression and activity in SOL, EDL and DIA and MMP2 mRNA expression in DIA. These results suggest that MMP changes may contribute to the skeletal muscle myopathy during heart failure.

Conjugação e validação de controle isotípico IgG1-FITC para uso em citometria de fluxo

Em meados da década de 50 iniciou-se o desenvolvimento da citometria de fluxo, tecnologia que permite verificar características físico-químicas de células ou partículas suspensas em meio fluido. Esta tecnologia utiliza anticorpos monoclonais marcados com fluorocromos como ferramenta de investigação em diversas análises e necessita de controles isotípicos para definição da região negativa (background). Estes controles são constituídos por imunoglobulinas de mesmo isotipo e fluorocromo dos anticorpos testes, sendo o isotiocianato de fluoresceína (FITC) o marcador fluorescente mais utilizado na conjugação de anticorpos. Os controles isotípicos têm como função definir a fluorescência inespecífica (células negativas) e as regiões fluorescentes (células positivas). No presente estudo foi selecionado anticorpo monoclonal murino (AcMm) dirigido contra antígeno eritrocitário canino, produzido no Laboratório de Anticorpos Monoclonais do Hemocentro de Botucatu, o qual reage positivamente com hemácias de cães, mas nunca com leucócitos humanos, tendo, portanto, potencial utilidade como controle negativo em citometria de fluxo. A purificação do AcMm da subclasse IgG1 foi feita por cromatografia de afinidade em Proteína-A Sepharose, e o controle da purificação realizado por eletroforese em géis de ágarose e poliacrilamida (SDS-PAGE). A imunoglobulina purificada foi conjugada ao FITC e filtrado em coluna de Sephadex G-25 para separação das proteínas marcadas e não-marcadas. O AcMm conjugado foi testado contra hemácias de cães, e o êxito da conjugação comprovado por testes de fluorescência, sendo a mediana de positividade de 94,70. Frente a leucócitos humanos a mediana de positividade foi 0,03 contra 0,50 dos reagentes comerciais. Os testes estatísticos não-paramétricos de Wilcoxon e correlação de Spearman comprovaram a eficiência e validam o controle isotípico produzido em comparação aos reagentes comerciais testados.

Genetic diversity of three size classes of seeds of Eucalyptus globulus ssp. globulus

Variation in seed size is often observed in samples of eucalypt seeds and this leads to heterogeneous populations of plants, principally through variation in the early stages of plant development. It follows that samples of seeds more uniform in size could produce more uniform populations of plants. In studies of Eucalyptus globulus ssp. globulus it was of interest to determine whether or not the genetic diversity within a population, through the use of isozyme markers, was altered in the subpopulations developed from seeds of different size classes. A commercial sample of seed was separated by seed size into three subpopulations and the percentage germination and mean fresh weight of the seedlings were determined. Proteins extracted from leaves of the seedlings were separated by electrophoresis and tested for activity of eight different enzymes. These eight enzymes showed activity at 20 loci and mean genetic diversity and fixation index were determined using 13 of these loci. The subpopulation of the smallest seeds contained a greater proportion of abnormal seeds and had a lower percentage germination and plant weight compared to the other subpopulations. No significant differences were found in the number of alleles per locus, percentage of polymorphic loci, mean heterozygosity. The major part of the endogamy, indicated by F statistic, was found within the subpopulations: F-(IS) = 0.518; F-(ST) = 0.010 and F(IT) = 0.523. We conclude that the use of seeds of uniform size will lead to more uniform germination and plant growth without alteration in overall genetic diversity.

Structural characteristics of the tendinous cord-papillary muscle junction in healthy and hypertensive rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Arabidopsis thaliana Uncoupling Proteins (AtUCPs): insights into gene expression during development and stress response and epigenetic regulation

Mitochondrial inner membrane uncoupling proteins (UCP) catalyze a proton conductance that dissipates the proton electrochemical gradient established by the respiratory chain, thus affecting the yield of ATP synthesis. UCPs are involved in mitochondrial energy flow regulation and have been implicated in oxidative stress tolerance. Based on the global gene expression profiling datasets available for Arabidopsis thaliana, in this review we discuss the regulation of UCP gene expression during development and in response to stress, and provide interesting insights on the possible existence of epigenetic regulation of UCP expression.

Interação entre Hb B2 e Hb S

As hemoglobinopatias e talassemias constituem as afecções genéticas mais comuns, apresentando-se, na maioria dos casos, em heterozigose. Diante da diversidade de hemoglobinas variantes encontrada na população brasileira, metodologias específicas e complementares para um diagnóstico laboratorial preciso, capaz de elucidar possíveis interações entre estas variantes genéticas, são necessárias. Este relato de caso descreve a interação entre hemoglobina B2 e a hemoglobina S em um indivíduo do sexo feminino, caucasoide, proveniente da região Sudeste do Brasil, identificada por meio de técnicas eletroforéticas em diferentes pH, cromatografia líquida de alta performance e PCR- RFLP. Visto que a hemoglobina B2 coelui com a hemoglobina S na análise cromatográfica e dificilmente é visualizada em eletroforese pH alcalino, devido à sua baixa concentração, justifica-se a necessidade da associação de testes laboratoriais, inclusive moleculares, na rotina do diagnóstico de hemoglobinas para a correta identificação do perfil de hemoglobinas do indivíduo e real frequência na população brasileira. Rev. Bras. Hematol. Hemoter.

Characterization and oxygen binding properties of des-Arg human hemoglobin

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Antidiarrheal mechanism of Carpolobia lutea leaf fractions in rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)