Avaliação do potencial genotóxico de nanopartículas de óxido de zinco e dióxido de titânio pelo ensaio do micronúcleo em células V79 e teste da mancha da asa em Drosophila melanogaster

The growing exploration of nanoparticles (NPs) engineered structures that have at least one dimension between 1 and 100 nm has a wide range of applications in many fields. The aim of the present study was evaluate the cytotoxic, mutagenic and recombinogenic effects of: 1] Amorphous Zinc oxide (ZnO), used as cement in endodontic; 2] ZnO NPs (21 nm); 3] three different types of Titanium dioxide (TiO2) NPs: anatase TiO2 (5.2 nm), anatase TiO2 (9.1 nm) and rutile TiO2 (55.1 nm). The tests employed were the in vivo wing Somatic Mutation and Recombination Test (SMART) in Drosophila melanogaster wings and the in vitro Cytokinesis-Block Micronucleus (CBMN) assay using Chinese hamster lung fibroblasts (V79 culture cells). D. melanogaster larvae of 72 ± 4 h, obtained from standard cross (ST) or high bioactivation cross (HB) were treated with 1.5625; 3.125; 6.25 or 12.5 mM to perform wing spot test (SMART). From both crosses, were obtained marker transheterozygous (MH) and balancer heterozygous (BH) descendants. The analysis of MH descendants showed that, for all composites, mutagenic and recombinogenic effects were observed only in HB cross. Statistically significant increase in the frequency of mutant spots was observed in individuals treated with 6.25 mM of amorphous ZnO and 12.5 mM of ZnO NPs, mainly due to induction of small single spots. The negative results observed in correspondent BH individuals, in witch is possible the detection only of mutation and chromosomal aberration, allow us to suggest that the increase in the frequency of small single spots observed in MH descendants is mainly due to recombination between flr3 and mwh loci. For TiO2 NPs, anatase of 5.2 nm was mutagenic at concentrations 1.5625 and 3.125 mM, and rutile phase of 55.1 nm significantly increased the frequency of mutant spots at all concentrations tested, except at 3.125 mM. Anatase TiO2 NPs (9.1 nm) did not cause mutagenic effects in wing of D. melanogaster neither in ST nor in HB crosses. The mutagenic effects observed in HB cross in contrast with ST cross in SMART assay can be justified possibly by the increase of metabolic activity in that cross, with an increasing production of reactive oxygen species and consequently higher DNA aggression. To determine the cell treatment for CBMN assay, several concentrations of ZnO NPs and three types of TiO2 NPs, ranging from 30.5 to 62,500.00 μM, were tested for cytotoxic activity by XTT colorimetric assay with xv V79 cells. ZnO NPs (21 nm) and anatase TiO2 NPs (5.2 and 9.1 nm) demonstrated similar cytotoxic intensity, showing decrease of V79 cell viability since concentration 224.1 μM. On the other hand, rutile TiO2 NPs (55.1 nm) cytotoxicity started later, at 7,812.00 μM. Among all NPs assessed, only ZnO (21 nm) and anatase TiO2 (5.2 nm) at highest concentration were able to induce genotoxicity in V79 cells. Both, anatase TiO2 NPs (9.1 nm) and rutile TiO2 NPs (55.1 nm) have not caused significant micronuclei increase. There are several physicochemical parameters that can influence on toxic effects of NPs, what corroborate to differences in cytotoxic and genotoxic patterns. In the experimental conditions performed in this study, we can affirm that both kinds of ZnO, as well, both, anatase and rutile phases of TiO2 NPs, were able to induce mutagenicity. Therefore, the use of these NPs should be closely monitored and its genotoxicity action mechanisms must be elucidated.Universidade de FrancaDoutor em Genética e BioquímicaA crescente exploração de nanopartículas estruturas sintetizadas artificialmente que apresentam pelo menos uma dimensão entre 1 e 100 nm está relacionada a vasta gama de aplicações em diferentes áreas. Devido às suas propriedades específicas, nanopartículas de óxido de zinco (NPs de ZnO) e nanopartículas de dióxido de titânio (NPs de TiO2) têm recebido muita atenção por causa das diversas possibilidade de utilização em produtos de consumo, principalmente em protetores solares, visto à eficácia aumentada na proteção contra luz UV e por serem menos opacas que os respectivos compostos de tamanho maior. O objetivo deste estudo foi avaliar os efeitos citotóxicos, mutagênicos e recombinogênicos de NPs de ZnO (21 nm), ZnO comercial, e três tipos diferentes de NPs de TiO2: TiO2 anatase (5,2 nm), TiO2 anatase (9,1 nm) e TiO2 rutila (55,1 nm). O teste in vivo Somatic Mutation and Recombination Test (SMART) em asa de Drosophila melanogaster e o teste do Micronúcleo com Bloqueio de Citocinese (CBMN), in vitro, utilizando fibroblastos de pulmão de hâmster Chinês (células V79) foram os escolhidos para realizar este trabalho. Larvas de 72 ± 4 h de D. melanogaster obtidas por meio do cruzamento padrão (ST) ou cruzamento de alta bioativação (HB) foram tratadas com 1,5625; 3,125; 6,25 ou 12,5 mM para o teste SMART. Para os dois tipos de compostos, ZnO e TiO2, foram observados efeitos mutagênicos apenas no cruzamento HB. Em relação ao ZnO, o comercial foi capaz de aumentar as frequências de manchas mutantes na concentração de 6,25 mM, enquanto NPs de ZnO induziram mutagenicidade na concentração mais alta. Para as NPs de TiO2, a fase anatase de 5,2 nm foi mutagênica nas concentrações 1,5625 e 3,125 mM, e a rutila de 55,1 nm aumentou de forma significativa a frequência de manchas mutantes em todas as concentrações avaliadas, exceto com 3,125 mM. NPs de anatase TiO2 (9,1 nm) não causaram efeitos mutagênicos em asas de D. melanogaster em nenhum dos cruzamentos. Os efeitos mutagênicos observados no cruzamento HB, em contraste com o cruzamento ST, no teste SMART podem ser justificados pela atividade metabólica aumentada inerente aos indivíduos HB, com aumento na produção de espécies reativas de oxigênio e, consequentemente, maior dano ao DNA. Para determinar as doses utilizadas no tratamento do teste do micronúcleo, várias concentrações xiii de NPs de ZnO, e dos três tipos de NPs de TiO2, variando de 30,5 a 62.500,00 μM, foram avaliadas pelo teste colorimétrico XTT em células V79. NPs de ZnO e TiO2 anatase de 5,2 e 9,1 nm demonstraram intensidade citotóxica similar, sendo que a partir da concentração 244,1 μM foi observada significativa redução da viabilidade celular. Por outro lado, a citotoxicidade de NPs de TiO2 na fase de rutila (55,1 nm) começou mais tardiamente, a partir a concentração de 7.812,00 μM. Dentre as NPs avaliados, apenas ZnO (21 nm) e TiO2 anatase (5,2 nm) na maior concentração avaliada foram capazes de induzir genotoxicidade em células V79. As fases de TiO2 anatase (9,1 nm) e TiO2 rutila (55,1 nm) não causaram aumento significativo de micronúcleo. Existem vários aspectos fisicoquímicos que podem influenciar nos efeitos tóxicos das NPs, o que corrobora com diferenças nos resultados citotóxicos e genotóxicos. Nas condicões experimentais deste estudo, podemos afirmar que ambos os tipos de ZnO avaliados, bem como as fases anatase e rutila das NPs de TiO2 foram capazes de induzir mutagenicidade. Portanto, o uso desses nanomateriais deve ser cuidadosamente monitorado e os mecanismos de indução de genotoxicidade devem ser elucidados

Novo marcador molecular para detecção e quantificação de Mycobacterium leprae por meio de PCR em tempo real

Leprosy epidemiologic studies have been restricted to Mycobacterium leprae DNA detection in nasal and bucal swabs with scarce literature on the peripheral blood, probably due to the difficulty in finding bacilli in this type of sample. We present the first molecular epidemiology study using real time PCR (qPCR) for M. leprae detection and quantification on peripheral blood of 200 leprosy patients and 826 household contacts, associating them with others clinical and laboratorial parameters. The TaqMan® qPCR system was performed with two groups of primers and probes, aiming the following M. leprae genomic regions: 85B antigen and ML0024. The marker 85B presented a low specificity, probably due to the unspecific DNA amplification from other microorganisms, while the new marker ML0024 described in this study, was 100% M. leprae specific, with a positivity of 22% in leprosy patients, but variable in the different clinical forms, and with significant increase in the bacillary load of lepromatous leprosy patients blood in relation to the tuberculoid form. The detection in household contacts was 1.2%. The presence of M. leprae DNA in the blood of contacts that developed leprosy presented an odds ratio of 17.22-fold higher in favor of the disease occurrence in relation to the healthy contacts. The present study describes the ML0024 as the most sensitive and specific marker for M. leprae DNA detection, and it is the largest molecular epidemiologic study in the peripheral blood of leprosy patients and their contacts, with profound implications on the disease management. The M. leprae detection on the blood of contacts has imposed an alert for the possible bacilli transmission through the blood and the probability of these healthy carriers in transmitting the bacilli to populations of regions where leprosy has been eliminated.Universidade de FrancaMestre em Ciências da SaúdeO estudo epidemiológico da hanseníase tem sido restrito à detecção do DNA de Mycobacterium leprae em swab nasal e pele, com escassa literatura em sangue periférico, provavelmente pela dificuldade de se encontrar o bacilo nesta amostra. Este é o primeiro trabalho de epidemiologia molecular que utilizou a PCR em tempo real (qPCR) para detectar e quantificar o DNA de M. leprae em sangue periférico de 200 pacientes com hanseníase e 826 contatos domiciliares, associando-os com os outros parâmetros clínico-laboratorais. A qPCR pelo sistema TaqMan® foi realizada com dois conjuntos de primers e sondas, tendo como alvos as regiões genômicas do antígeno 85B e ML0024 do bacilo. O marcador 85B não foi específico, pois amplificou DNA de outros parasitos, enquanto que o novo marcador ML0024, descrito neste trabalho, foi 100% específico ao DNA do bacilo, com uma positividade de 22% nos pacientes, porém variável nas diversas formas clínicas e com significante aumento da quantificação da carga bacilar no sangue de pacientes V em relação aos T. Nos contatos a detecção foi de 1,2%. A presença do DNA de M. leprae em sangue de contatos que desenvolveram a doença revelou uma chance 17.22 vezes maior de adoecer em relação aos contatos sadios. Este trabalho descreveu o marcador ML0024 como o mais sensível e específico para o M. leprae e realizou um amplo estudo em sangue periférico na hanseníase, que pode ter implicações na epidemiologia da doença. A detecção de M. leprae no sangue de contatos, considerados portadores sadios, alerta para a transmissão sanguínea do bacilo e para a possibilidade desses indivíduos serem carreadores do bacilo para regiões onde a hanseníase já foi eliminada

Evaluation of Compressive Strength of Manganese Oxidized Ore Briquettes: Effects of Compaction Pressure, Curing Time, and Percentage of Binder

Agglomeration is a common process to safely recycle wastes from the mining and metallurgical sectors. In this process, the mineralogical and chemical characteristics of manganese ores exert a significant influence on the reduction reactions within the electric furnace. Therefore, a systematic work has been done to evaluate the compressive strength of briquettes composed of manganese oxidized ore fines, ferromanganese dust, charcoal fines, and three different binders, bentonite, pregelatinized starch, and slaked lime + molasses. A response surface methodology was used to evaluate the effects of compaction pressure, curing time, and % binder, whose response was measured in terms of compressive strength. The results were assessed by Analysis of Variance and a regression model was generated. The study showed that briquettes with bentonite demonstrated an optimized compressive strength of 2.54 MPa with a compaction pressure of 52.5 MPa, 11 days of curing time, and 11.5% bentonite content. Briquettes with slaked lime + molasses achieved a compressive strength of 3.70 MPa with a compaction pressure of 62.12 MPa, a curing time of 24 days, and an addition of 16.2% slaked lime + molasses. For briquettes produced with pre-gelatinized starch, the design of the proposed variables resulted in a lack of fit to the model. Optimization studies revealed that briquettes made with bentonite and slaked lime + molasses are viable alternatives to replace oxidized lump ore in electric furnaces.</p

Molecular diagnosis of canine visceral leishmaniasis : a comparative study of three methods using skin and spleen from dogs with natural Leishmania infantum infection.

Polymerase chain reaction (PCR) and its variations represent highly sensitive and specificmethods for Leishmania DNA detection and subsequent canine visceral leishmaniasis (CVL)diagnosis. The aim of this work was to compare three different molecular diagnosis tech-niques (conventional PCR [cPCR], seminested PCR [snPCR], and quantitative PCR [qPCR])in samples of skin and spleen from 60 seropositive dogs by immunofluorescence antibodytest and enzyme-linked immunosorbent assay. Parasitological analysis was conducted byculture of bone marrow aspirate and optical microscopic assessment of ear skin and spleensamples stained with Giemsa, the standard tests for CVL diagnosis. The primers L150/L152and LINR4/LIN17/LIN19 were used to amplify the conserved region of the Leishmania kDNAminicircle in the cPCR, and snPCR and qPCR were performed using the DNA polymerasegene (DNA pol _) primers from Leishmania infantum. The parasitological analysis revealedparasites in 61.7% of the samples. Sensitivities were 89.2%, 86.5%, and 97.3% in the skin and81.1%, 94.6%, and 100.0% in spleen samples used for cPCR, snPCR, and qPCR, respectively.We demonstrated that the qPCR method was the best technique to detect L. infantum inboth skin and spleen samples. However, we recommend the use of skin due to the highsensitivity and sampling being less invasive

Unveiling healthy carriers and subclinical infections among household contacts of leprosy patients who play potential roles in the disease chain of transmission

Leprosy transmission still occurs despite the availability of highly effective treatment. The next step towards successfully eliminating leprosy is interrupting the chain of transmission of the aetiological agent, Mycobacterium leprae. In this investigation, we provide evidence that household contacts (HHCs) of leprosy patients might not only have subclinical infections, but may also be actively involved in bacilli transmission. We studied 444 patients and 1,352 contacts using anti-phenolic glycolipid-I (PGL-I) serology and quantitative polymerase chain reaction (qPCR) to test for M. leprae DNA in nasal swabs. We classified the patients according to the clinical form of their disease and the contacts according to the characteristics of their index case. Overall, 63.3% and 34.2% of patients tested positive by ELISA and PCR, respectively. For HHCs, 13.3% had a positive ELISA test result and 4.7% had a positive PCR test result. The presence of circulating anti-PGL-I among healthy contacts (with or without a positive PCR test result from nasal swabs) was considered to indicate a subclinical infection. DNA detected in nasal swabs also indicates the presence of bacilli at the site of transmission and bacterial entrance. We suggest that the concomitant use of both assays may allow us to detect subclinical infection in HHCs and to identify possible bacilli carriers who may transmit and disseminate disease in endemic regions. Chemoprophylaxis of these contacts is suggested

Painel 2 - Experiências Setoriais: o Monitoramento nas Áreas da Educação e Saúde

Painel 2 - Experiências Setoriais: o Monitoramento nas Áreas da Educação e Saúde; Palestra 1 - Ministério da Educação: "Instrumentos para Monitoramento e Avaliação das Políticas e Programas do Ministério da Educação", apresentada por Milena Lins Fernandes Soares e Érica de Oliveira; Palestra 2 - Ministério da Saúde: "O Monitoramento e a Avaliação na Gestão do Ministério da Saúde", apresentada por Afonso Teixeira dos ReisApresentação de slides em PDF - Painel 2.Políticas Públicas e Sociai

Biodegradation of Poly (3-hydroxybutyrate) /Eggshellsystems

<div><p>In this work, biocomposites of poly (3-hydroxybutyrate) (PHB) / calcium carbonate from Rhea Americana eggshells were prepared and the effects of the addition of the inorganic filler in the polymeric matrix were assessed. The residue (powder) of the eggshell calcined at 400 ºC or in natura was inserted into a PHB solution for preparation of films via casting. Powder samples were characterized by X-Ray Fluorescence (XRF), X-Ray diffraction (XRD) and Thermogravimetry (TG/DTG) and films were characterized by X-Ray diffraction (XRD), Scanning Electron Microscopy (SEM) and biodegradation tests according to the ASTM G 160-03 norm: the results were reported as weight loss and visual inspection by optical microscopy (OM). From the results of the XDR, it was perceived that the peaks in the diffractograms of the powder obtained by milling the Rhea Americana eggshells corresponded to the diffraction patterns of the Calcite crystals, which is a calcium carbonate polyform, and that the calcination preserved the crystalline structure, maintaining the calcium carbonate in the samples. For the biocomposites, a peak characteristic to the calcium carbonate in 29.57º was detected, indicating the insertion of the filler to the polymer matrix. Through SEM the presence of small agglomerates, probably due to polymer particles that were not dissolved, was observed for the pure PHB film. With the addition of the filler in natura a greater porosity was formed in the surface of the biocomposite films, and with the calcined filler, homogeneous films with reduced porosity were obtained. From the weight loss and OM results, it was observed that the filler inserted into the polymeric matrix catalyzes the biodegradation process up to 60 days evaluation in different ways, depending on the type of sample used.</p></div