Análise do potencial microbiano de uma biopilha na biorremediação de solos contaminados por hidrocarbonetos de petróleo / Analysis of microbial potential of a biopile in soil bioremediation contaminated by petroleum hydrocarbons

RESUMO A presente pesquisa avaliou o potencial microbiano de uma biopilha na biorremediação de solos contaminados por hidrocarbonetos, montada em uma base de distribuição de combustíveis na região metropolitana de Porto Alegre, Rio Grande do Sul. Desta biopilha, foram avaliadas as concentrações dos hidrocarbonetos totais de petróleo (HTP) e de compostos benzeno, tolueno, etilbenzeno e xilenos (BTEX), em nove pontos, durante quatro etapas de operação e em três profundidades por ponto. De cada coleta, também foram reservadas amostras de solo para realização das análises microbiológicas. A partir dessas amostras, buscou-se identificar fungos e leveduras pela análise de suas estruturas reprodutivas em lâminas de microcultivo, e bactérias pela técnica da Reação em Cadeira da Polimerase (PCR) e sequenciamento do gene que codifica o RNAr 16S. Ainda, avaliou-se a capacidade dos microrganismos isolados em degradar óleo diesel comercial, utilizando o indicador redox 2,6-diclorofenol indofenol (DCPIP). Como resultado deste estudo, observaram-se elevados percentuais de redução nas concentrações de HTP e BTEX na biopilha, como 92 e 100%, respectivamente. Das amostras de solo da biopilha, foram isoladas 101 cepas de microrganismos, das quais foram identificadas 19 cepas de fungos filamentosos, 34 de bactérias e 1 de levedura. Os resultados evidenciaram a capacidade de alguns gêneros de fungos, como Aspergillus, Trichoderma, Penicillium, Cladosporium e Verticillium, e bactérias, como Bacillus spp. e Streptomyces sp., em degradar hidrocarbonetos constituintes do óleo diesel comercial.

ABSTRACT This research evaluated the microbial potential of a biopile in bioremediation of soils contaminated by hydrocarbons, mounted on a fuel distribution base in the metropolitan region of Porto Alegre, Rio Grande do Sul. Of this biopile were evaluated concentrations of total petroleum hydrocarbons (TPH) and benzene compounds, toluene, ethylbenzene and xylenes (BTEX) at nine points, for four stages of operation and a point in three depths. Of each collection were also reserved sampled soil to perform the microbiological testing. From the samples, we sought to identify fungi and yeasts by analyzing their reproductive structures microcultivation blades, and bacteria by the technique of Polymerase Reaction Chair (PCR) and gene sequencing encoding the 16S rRNA. Still, we evaluated the ability of microorganisms to degrade commercial diesel oil, using the redox indicator 2,6-dichlorophenol indophenol (DCPIP). As result of this study, there was a high percentage reduction in the concentration of TPH and BTEX in biopilha as 92 and 100%, respectively. Of soil samples were isolated from biopilha 101 strains of microorganisms, of which 19 were identified strains of filamentous fungi and 34 bacterial yeast. The results showed the ability of some genera of fungi such as Aspergillus, Trichoderma, Penicillium, Cladosporium and Verticillium and bacteria such as Bacillus spp. and Streptomyces sp. to degrade hydrocarbons constituents of commercial diesel oil.

Quantification of glucosylceramide in plasma of Gaucher disease patients

Gaucher disease is a sphingolipidosis that leads to an accumulation of glucosylceramide. The objective of this study was to develop a methodology, based on the extraction, purification and quantification of glucosylceramide from blood plasma, for use in clinical research laboratories. Comparison of the glucosylceramide content in plasma from Gaucher disease patients, submitted to enzyme replacement therapy or otherwise, against that from normal individuals was also carried out. The glucosylceramide, separated from other glycosphingolipids by high performance thin layer chromatography (HPTLC) was chemically developed (CuSO4 / H3PO4) and the respective band confirmed by immunostaining (human anti-glucosylceramide antibody / peroxidase-conjugated secondary antibody). Chromatogram quantification by densitometry demonstrated that the glucosylceramide content in Gaucher disease patients was seventeen times higher than that in normal individuals, and seven times higher than that in patients on enzyme replacement therapy. The results obtained indicate that the methodology established can be used in complementary diagnosis and for treatment monitoring of Gaucher disease patients.

A doença de Gaucher é uma esfingolipidose caracterizada pelo acúmulo de glicosilceramida. O objetivo deste estudo foi desenvolver metodologia baseada na extração, purificação e quantificação da glicosilceramida plasmática a qual possa ser usada em laboratórios de pesquisa clínica. Após o desenvolvimento desta metodologia, foi proposto, também, comparar o conteúdo de glicosilceramida presente no plasma de pacientes com doença de Gaucher, submetidos ou não a tratamento, com aquele de indivíduos normais. A glicosilceramida, separada de outros glicoesfingolipídios por cromatografia de camada delgada de alto desempenho (HPTLC), foi revelada quimicamente (CuSO4/H3PO4) e a respectiva banda foi confirmada por imunorrevelação (anticorpo anti-glicosilceramida humana/anticorpo secundário conjudado à peroxidase). A quantificação do cromatograma por densitometria demonstrou que o conteúdo de glicosilceramida nos pacientes com doença de Gaucher era 17 vezes maior que aquele de indivíduos normais e 7 vezes maior que aquele dos pacientes com doença de Gaucher submetidos a tratamento com terapia de reposição enzimática. Os resultados obtidos demonstram que a metodologia estabelecida pode ser usada como diagnóstico complementar e como monitoração do tratamento de pacientes com doença de Gaucher.

The use of RAPD to characterize Bipolaris sorokiniana isolates.

Bipolaris sorokiniana is a phytopathogenic fungus causing diseases of cereal crops such as common root rot, the leaf spot disease, seedling blight, and black point of the grain. Random-amplified polymorphic DNA (RAPD) assay was used to investigate the genetic diversity of 20 isolates collected from different cultivars in wheat-producing regions in Brazil. Seventy primers, with random nucleotide sequences, were tested. Reproducibility to amplify the genomic DNA of isolates was found for 30 of the 70 primers tested, generating between 1 and 17 fragments ranging from 0.35 to 2.0 kb (average size). The degree of similarity between samples was calculated through simple association and the dendrogram was assessed using the unweighted pair group method with arithmetical average. After the RAPD analyses 19 isolates were closely grouped, having a similarity coefficient of >or= 78%. Isolate I017 showed very low similarity coefficients, ranging between 38 and 46%. The RAPD analyses provided important information as to the degree of genetic variability and the relationship between the isolates investigated, revealing polymorphism and establishing electrophoretic profiles useful to characterize the phytopathogen.