Remoção de herbicida atrazina por meio de filtros de carvão ativado granular associados com microrganismos no tratamento de água para abastecimento / Removal of atrazine herbicide through granular activated carbon filters associated with microorganisms in drinking water treatment

RESUMO A atrazina é um herbicida sintético comumente utilizado no controle de ervas gramíneas daninhas e folhagens em lavouras, e é um dos principais contaminantes dos solos e dos ecossistemas aquáticos. Muitos métodos têm sido sugeridos para remover herbicidas da água potável. Contudo, esses métodos são custosos, muitos têm problemas de desempenho, produzem diversos subprodutos intermediários tóxicos, prejudiciais e perigosos. Entretanto, a atrazina é susceptível à degradação por microrganismos presentes na água, no sedimento e no esgoto. Considerando esses aspectos, o objetivo principal do estudo foi investigar a biodegradação e a filtração por meio de filtros de carvão com atividade biológica (CAB) para remoção da atrazina, e sua identificação filogenética associada a esses microrganismos. Os resultados demonstraram que a atrazina foi degradada por microrganismos presentes no biofilme, com remoção superior a 89% nos filtros CAB. Os microrganismos encontrados integram-se ao grupo das bactérias, composto dos gêneros Acinetobacter,Bacillus, Exiguobacterium e Pseudomonas. Este estudo nos permite inferir sobre a capacidade de biodegradação da atrazina por bactérias presentes nos filtros CAB, a capacidade de remover herbicidas por meio desse sistema de filtros e a possível utilização dessa tecnologia como alternativa para o controle e a remoção dessa substância no tratamento de água.

ABSTRACT Atrazine is a synthetic herbicide commonly used to control weeds and foliage in crops, and is a major contaminants of soil and water ecosystems. Many methods have been suggested to remove herbicides from drinking water. However, these methods are very costly, many have performance problems, produce a lot of toxic intermediates which are very harmful and dangerous. However, atrazine is susceptible to degradation by microorganism present in water, sediment, and sewage effluents. Considering these aspects, the main objective of the study was to investigate the biodegradation and filtration for using biological activated carbon (BAC) filters to remove atrazine, and their phylogenetic identification associated with these microorganisms. The results showed that atrazine was biodegraded by microorganism present in the biofilm, with removal over 80% in BAC filters. The microorganisms found integrate to the group of bacteria, composed by the genera Acinetobacter, Bacillus, Exiguobacterium, and Pseudomonas. This study allows us to infer the ability to biodegrade atrazine by bacteria present in BAC filters and capacity to remove herbicides by BAC filters, and the possible use of this technology as an alternative for the control and removal of this substance in water treatment.

Bacterial communities in mining soils and surrounding areas under regeneration process in a former ore mine

Abstract Human activities on the Earth's surface change the landscape of natural ecosystems. Mining practices are one of the most severe human activities, drastically altering the chemical, physical and biological properties of the soil environment. Bacterial communities in soil play an important role in the maintenance of ecological relationships. This work shows bacterial diversity, metabolic repertoire and physiological behavior in five ecosystems samples with different levels of impact. These ecosystems belong to a historical area in Iron Quadrangle, Minas Gerais, Brazil, which suffered mining activities until its total depletion without recovery since today. The results revealed Proteobacteria as the most predominant phylum followed by Acidobacteria, Verrucomicrobia, Planctomycetes, and Bacteroidetes. Soils that have not undergone anthropological actions exhibit an increase ability to degrade carbon sources. The richest soil with the high diversity was found in ecosystems that have suffered anthropogenic action. Our study shows profile of diversity inferring metabolic profile, which may elucidate the mechanisms underlying changes in community structure in situ mining sites in Brazil. Our data comes from contributing to know the bacterial diversity, relationship between these bacteria and can explore strategies for natural bioremediation in mining areas or adjacent areas under regeneration process in iron mining areas.

Carbohydrates on the growth of Cattleya schilleriana Reichb.f. seedlings (Orchidaceae) / Carboidratos no crescimento de plântulas de Cattleya schilleriana Reichb.f. (Orchidaceae)

ABSTRACT: The aim of the present study was to evaluate the effects of carbohydrate supplementation on the propagation of the orchid Cattleya schilleriana. The 120-d-old seedlings were subcultured in fructose-, glucose-, or sucrose-supplemented (0, 15, 30, and 45g L-1) ½ MS culture medium (half-strength macronutrient concentrations), using a completely random design with four repetitions per treatment. After 120d of treatment, root number and length, leaf number and length, and fresh weight were evaluated, and seedling survival was evaluated after 75d of acclimatization in a greenhouse. The in vitro growth data were submitted to regression analysis, whereas the percentage survival data were analyzed using ANOVA and Tukey's test. Both in vitro growth and ex vitro survival were lowest when the plantlets were grown in the absence of a carbohydrate source and highest (>90% survival) when supplemented with glucose. According to our findings, the addition of either glucose (30g L-1) or sucrose (30g L-1) is recommended for mass propagation of C. schilleriana.

RESUMO: O objetivo deste estudo foi avaliar a adição de doses de carboidratos em meio de cultura ½ MS para a propagação da orquídea Cattleya schilleriana. Plântulas com 120 dias após semeadura foram recultivadas em 12 tratamentos, onde foram investigadas a adição de diferentes doses de frutose, glicose e sacarose (0, 15, 30 e 45g L-1). O delineamento utilizado foi inteiramente casualizado com quatro repetições por tratamento. Após 120 dias nos tratamentos, foram mensurados o número de raízes, comprimento da maior raiz, número de folhas, comprimento da maior folha e massa fresca total. Depois as plântulas foram aclimatizadas em casa de vegetação e avaliada a taxa de sobrevivência após 75 dias. Os dados obtidos foram submetidos ao teste de Tukey a 5% de probabilidade e ajustadas curvas de regressão. A ausência de carboidrato reduziu o crescimento das plântulas entre todas as características avaliadas in vitro e ex vitro. As doses de glicose promoveram maior eficiência para o crescimento in vitro e sobrevivência acima de 90% após a aclimatização. Assim, a adição de glicose (30g L-1) ou sacarose (30g L-1) são recomendadas para a propagação massal de C. schilleriana.

Uso de filtros de carvão ativado granular associado a microrganismos para remoção de fármacos no tratamento de água de abastecimento / Use of granular activated carbon filters associated with microorganisms to remove pharmaceuticals in drinking water treatment

RESUMO O modo de vida das sociedades modernas originou o aporte diário, nos ambientes aquáticos, de fármacos e outras inúmeras moléculas de uso contínuo, compostos emergentes, com potencial de risco à saúde humana principalmente pela exposição em razão da inevitável contaminação dos mananciais de abastecimento de água e da transferência para as estações de tratamento de água (ETA), onde não são removidos. O uso de carvão ativado granular na filtração demonstra ser uma opção viável para ETA, porém, uma eficiência satisfatória requer regeneração periódica do material, onerando o tratamento. Contudo, observa-se que em baixas taxas de filtração a colonização natural dos filtros por microrganismos - formação de biofilme - pode ser uma alternativa para aumentar o tempo de vida útil do carvão, bem como para decompor essas moléculas complexas em elementos minerais assimiláveis, reintroduzindo-os nos ciclos biogeoquímicos naturais. Este trabalho avaliou, durante 24 semanas, em condições de laboratório, o carvão ativado com biofilme como meio filtrante para remoção dos fármacos: diclofenaco de sódio, ibuprofeno, naproxeno e amoxicilina; experimentou em sistema batch o potencial dos microrganismos colonizadores de filtros em degradar os fármacos testados, assim como identificou filogeneticamente os microrganismos predominantes na biodegradação. Os resultados demonstram a remoção dos fármacos acima de 80%. Constatou-se a presença das bactérias dos gêneros Bacillus , Burkholderia , Cupriavidus , Pseudomonas , Shinella , e Sphingomonas . Este estudo permite inferir a capacidade de remoção de fármacos por bactérias presentes em filtros de carvão ativado e o possível uso dessa tecnologia como alternativa de controle e remoção dessas substâncias no tratamento de água potável.

ABSTRACT The way of life of modern societies has originated the daily intake of pharmaceuticals and numerous other molecules of continuous use in aquatic environments, emerging compounds that brings potential risk for human health mainly due to exposure resulted from the inevitable contamination of sources of drinking water supply and its transference to the water treatment plants (WTP) where they are not removed. The use of granular activated carbon in filters proves to be a viable option for WTP, but satisfactory efficiency requires periodic regeneration of the material, burdening the treatment costs. However, it is noted that under low filtration rates, the natural colonization of filters by microorganisms - biofilm formation - may be an alternative for increasing the lifetime of carbon, as well as to decompose these complex molecules into assimilable mineral elements, thereby reintroducing them to the natural biogeochemical cycles. This study evaluated the activated carbon with biofilm as the filter media, during 24 weeks, under laboratory conditions, considering the removal of the pharmaceuticals diclofenac sodium, ibuprofen, naproxen and amoxicillin; experienced under batch system the potential of the microorganisms adhering to the filters in degrade the tested drugs, as well as phylogenetically identified the predominant microorganisms in biodegradation. The results show drug removal over 80%. It was observed the presence of the bacteria genus Bacillus, Burkholderia, Cupriavidus, Pseudomonas, Shinella and Sphingomonas. This study allows us to infer the capacity to remove pharmaceuticals by the bacteria present in the activated carbon filters, and the possible use of this technology as an alternative for control and removal of these substances in drinking water treatment.

Biotechnology of polyketides: new breath of life for the novel antibiotic genetic pathways discovery through metagenomics

The discovery of secondary metabolites produced by microorganisms (e.g., penicillin in 1928) and the beginning of their industrial application (1940) opened new doors to what has been the main medication source for the treatment of infectious diseases and tumors. In fact, approximately 80 years after the discovery of the first antibiotic compound, and despite all of the warnings about the failure of the "goose that laid the golden egg," the potential of this wealth is still inexorable: simply adjust the focus from "micro" to "nano", that means changing the look from microorganisms to nanograms of DNA. Then, the search for new drugs, driven by genetic engineering combined with metagenomic strategies, shows us a way to bypass the barriers imposed by methodologies limited to isolation and culturing. However, we are far from solving the problem of supplying new molecules that are effective against the plasticity of multi- or pan-drug-resistant pathogens. Although the first advances in genetic engineering date back to 1990, there is still a lack of high-throughput methods to speed up the screening of new genes and design new molecules by recombination of pathways. In addition, it is necessary an increase in the variety of heterologous hosts and improvements throughout the full drug discovery pipeline. Among numerous studies focused on this subject, those on polyketide antibiotics stand out for the large technical-scientific efforts that established novel solutions for the transfer/engineering of major metabolic pathways using transposons and other episomes, overcoming one of the main methodological constraints for the heterologous expression of major pathways. In silico prediction analysis of three-dimensional enzymatic structures and advances in sequencing technologies have expanded access to the metabolic potential of microorganisms.

Biodegradacao da hepatotoxina(D-Leu1)-microcistina-LR por bacterias presentes em filtros biologicos de carvao / Biodegradation of hepatotoxin (D-Leu1)-microcystin-LR by bacteria in carbon biological filters

A persistência das microcistinas (MCs) em ambientes aquáticos e sua difícil remoção no tratamento convencional de água representam um desafio às companhias de saneamento. Contudo, as MCs são susceptíveis à degradação por bactérias presentes na água, sedimentos e efluentes de esgotos. Neste estudo, avaliou-se a biodegradação de MCs por microrganismos presentes em filtros de carvão com atividade biológica (CAB) e sua identificação filogenética pelo sequenciamento do gene 16S RNA. Foi utilizada uma água de estudo contendo MCs com diferentes composições, acrescida de efluente de filtros CAB. Os resultados demonstraram que as MCs foram biodegradadas por microrganismos presentes no biofilme. Este estudo infere sobre a capacidade de biodegradação de MCs por bactérias presentes em filtros CAB e o possível uso destes microorganismos como alternativa de remoção de MCs no tratamento de água potável.

The persistence of MCs in aquatic environments and their difficult removal in the conventional water treatment is a challenge to companies of sanitation. However, the MCs are susceptible to degradation by bacteria present in water, sediment and sewage effluents. In this study, we investigated the biodegradation of MCs by microorganism present in carbon filters with biological activity (BAC) and their phylogenetic identification by sequencing gene 16S RNA. A study of water containing MCs was used, with different compositions, plus a filters BAC effluent. The results showed that of MCs were biodegraded by microorganism present in the biofilm. This study provides the ability to complete biodegradation of MCs by bacteria present in BAC filters and the possible use of these microorganisms as alternative of the removal of MCs in the treatment of drinking water.

Desenvolvimento inicial e crescimento in vitro de Cattleya violacea (Kunth) Rolfe em diferentes concentrações de sacarose / Initial development and in vitro growth of Cattleya violacea (Kunth) Rolfe in different sucrose concentrations

The aims of this study were to analyze the initial germination aspects and evaluate the effect of sucrose concentrations on in vitro growth of Cattleya violacea. Mature seeds from closed pods were sowed on MS (Murashige and Skoog) culture medium. The external morphology of seed and plantlets were documented by light and scanning electron microscopy. Plantlets with 90 days after sowing were subcultured on ½ MS (with half-strength macronutrients concentrations) culture medium with different sucrose concentrations (0, 10, 20, 30 and 40 g L-1), incubated under the same in vitro conditions during more 150 days followed by the evaluation of root number, root length, shoot length, fresh and dry weight. The biometric data were statistically analyzed and regression curves constructed. The seeds showed reticulate seed coat with a micropylar (opened) and chalazal (closed) end; the embryo results a chlorophyllated tuberiforme structure called protocorm, which may have rhizoids, leaflets and is considered plantlet when it has root. Absence of sucrose or the highest evaluated sucrose concentration was detrimental for plant growth. The concentration of 27 g L-1 provided the highest in vitro growth enabling great efficiency for mass propagation of this species of high ornamental potential.

O presente estudo teve por objetivo analisar os aspectos da germinação e avaliar o efeito de concentrações de sacarose no crescimento in vitro de Cattleya violacea. Sementes provenientes de cápsulas fechadas foram semeadas em meio de cultura Murashige e Skoog (MS) e a morfologia externa da semente à plântula foi fotodocumentada em estereomicroscópio e microscópio eletrônico de varredura. Plântulas com 90 dias após a semeadura foram repicadas em meio de cultura ½ MS (com metade da concentração de macronutrientes) com diferentes concentrações de sacarose (0, 10, 20, 30 e 40 g L-1), incubadas nas mesmas condições in vitro por mais 150 dias e em seguida as plântulas foram avaliadas quanto ao número de raízes, comprimento da maior raiz, número de folhas, comprimento da parte aérea, massa fresca e seca total. Os dados biométricos foram submetidos à análise estatística e a eles ajustadas curvas de regressão. As sementes apresentaram testa reticulada com uma extremidade micropilar (aberta) e calazal (fechada); o embrião originou uma estrutura tuberiforme clorofilada denominada protocormo que pode apresentar rizóides, folíolos e quando provido de raiz é considerado plântula. A ausência de açúcar ou a maior concentração avaliada de sacarose foram prejudiciais ao crescimento da planta. A concentração de 27 g L-1 proporcionou maior crescimento in vitro possibilitando maior eficiência para a propagação massal dessa espécie de elevado potencial ornamental.

Crescimento in vitro e aclimatização de Cattleya loddigesii Lindley (Orchidaceae) com carvão ativado sob dois espectros luminosos / In vitro growth and acclimatization of Cattleya loddigesii Lindley (Orchidaceae) with actived charcoal in two light spectra

O objetivo deste estudo foi avaliar a adição de concentrações de carvão ativado em meio de cultura ½ MS (com metade da concentração dos macronutrientes) sob dois espectros luminosos para a obtenção de plântulas in vitro de Cattleya loddigesii. Plântulas com aproximadamente 90 dias foram subcultivadas em oito tratamentos, nos quais foi testada a adição ao meio de cultura ½ MS com carvão ativado (0; 0,5; 1,0 e 2,0g L-1) e combinados sob espectro de luz branca e luz vermelha. Após 180 dias da germinação, foram mensurados dados biométricos (raiz e parte aérea), massa fresca e teores de pigmentos fotossintéticos. Em plântulas aclimatizadas em casa de vegetação, foram avaliadas a taxa de sobrevivência após 120 dias. As concentrações de clorofila total, clorofila a e carotenoides foram maiores nos tratamentos sob luz branca, enquanto a luz vermelha influenciou significativamente maior clorofila b, plântulas com menos raízes e de menor comprimento e elevada mortalidade ex vitro. A adição de 2,0g L-1 de carvão ativado ao meio de cultura e o uso de luz branca proporcionaram maior eficiência de desenvolvimento tanto para as culturas in vitro quanto para a sobrevivência ex vitro das plantas.

The aim of this study was to evaluate the effect of different concentrations of activated charcoal in ½ MS (half concentration of macronutrients) culture medium under two light spectra on the in vitro growth of Cattleya loddigesii seedlings. Plantlets with approximately 90 days were subcultured under eight treatments, consisting of different active charcoal concentrations (0; 0.5; 1.0 and 2.0g L-1) in ½ MS medium combined with white and red light spectra. After 180 days of germination, biometric data, fresh weight, and the level of photosynthetic pigments were evaluated. Plantlets acclimatized in a greenhouse were evaluated for survival after 120 days. Total chlorophyll, chlorophyll a, and carotenoid concentrations were higher in treatments under white light, while red light promoted greater chlorophyll b, plantlets with fewer and shorter roots, and high ex vitro mortality. The addition of 2.0g L-1 of active charcoal to the culture medium and the use of white light provided greater development efficiency both on in vitro culture and ex vitro plant survival.

Production of DNA microarray and expression analysis of genes from Xylella fastidiosa in different culture media

DNA Microarray was developed to monitor the expression of many genes from Xylella fastidiosa, allowing the side by-side comparison of two situations in a single experiment. The experiments were performed using X. fastidiosa cells grown in two culture media: BCYE and XDM2. The primers were synthesized, spotted onto glass slides and the array was hybridized against fluorescently labeled cDNAs. The emitted signals were quantified, normalized and the data were statistically analyzed to verify the differentially expressed genes. According to the data, 104 genes were differentially expressed in XDM2 and 30 genes in BCYE media. The present study showed that DNA microarray technique efficiently differentiate the expressed genes under different conditions.

DNA Microarray foi desenvolvida para monitorar a expressão de muitos genes de Xylella fastidiosa, permitindo a comparação de duas situações distintas em um único experimento. Os experimentos foram feitos utilizando células de X. fastidiosa cultivada em dois meios de cultura: BCYE e XDM2. Pares de oligonucleotídeos iniciadores foram sintetizados, depositados em lâminas de vidro e o arranjo foi hibridizado contra cDNAs marcados fluorescentemente. Os sinais emitidos foram quantificados, normalizados e os dados foram estatisticamente analisados para verificar os genes diferencialmente expressos. De acordo com nossos dados, 104 genes foram diferencialmente expressos para o meio de cultura XDM2 e 30 genes para o BCYE. No presente estudo, nós demonstramos que a técnica de DNA microarrays eficientemente diferencia genes expressos sob diferentes condições de cultivo.

Molecular characterization of bacterial populations of different soils

Until recently, few studies were carried out in Brazil about diversity of bacterial soil communities. Aiming to characterize the bacterial population in the soil through 16S rRNA analysis, two types of soil have been analyzed: one of them characterized by intensive use where tomato, beans and corn were cultivated (CS); the other analyzed soil was under forest (FS), unchanged by man; both located in Guaíra, São Paulo State, Brazil. Using specific primers, 16S rRNA genes from metagenomic DNA in both soils were amplified by PCR, amplicons were cloned and 139 clones from two libraries were partially sequenced. The use of 16S rRNA analysis allowed identification of several bacterial populations in the soil belonging to the following phyla: Acidobacteria, Actinobacteria, Bacteroidetes, Firmicutes, Proteobacteria Verrucomicrobia in addition to the others that were not classified, beyond Archaea domain. Differences between FS and CS libraries were observed in size phyla. A larger number of phyla and, consequently, a greater bacterial diversity were found in the under-forest soil. These data were confirmed by the analyses of genetic diversity that have been carried out. The characterization of bacterial communities of soil has made its contribution by providing facts for further studies on the dynamics of bacterial populations in different soil conditions in Brazil.

Até o momento poucos estudos foram realizados no Brasil a respeito da diversidade de comunidades bacterianas no solo. Com o objetivo de caracterizar as populações bacterianas presentes no solo através da análise do gene 16S rRNA, foram analisados dois solos: um caracterizado pelo uso intensivo, principalmente para a produção de tomate, feijão e milho (CS); e outro sob floresta (FS), não modificado pelo homem, ambos do município de Guaíra, no estado de São Paulo, Brasil. Usando oligonucleotídeos específicos, de genes 16S rRNA do DNA metagenomico de ambos os solos foram amplificados por PCR, amplicons foram clonados e 139 clones de duas bibliotecas foram seqüenciados. O uso da técnica de 16S rRNA, gerou a identificação de diferentes populações de bactérias de solo pertencentes aos filos Acidobacteria Actinobacteria Bacteroidetes Firmicutes Proteobacteria Verrucomicrobia, Archaea, além das não classificadas. Diferenças entre as bibliotecas FS e CS foram observadas no tamanho dos filos. Um grande número de filos e, consequentemente, uma grande diversidade bacteriana foi observada no solo sob floresta. Estes dados foram confirmados pela análise de diversidade genética realizada. A caracterização de comunidades do solo apresentada neste trabalho contribuiu fornecendo dados para estudos posteriores sobre a dinâmica das populações bacterianas em solos de diferentes condições no Brasil.

Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

The objective of this work was to assess the functionality of the glycolytic pathways in the bacterium Xylella fastidiosa. To this effect, the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase of the glycolytic pathway, and glucose 6-phosphate dehydrogenase of the Entner-Doudoroff pathway were studied, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates, which explains the increased duplication time of this phytopatogen. Recombinant enolase was expressed as inclusion bodies and solubilized with urea (most efficient extractor), Triton X-100, and TCA. Enolase extracted from X. fastidiosa and from chicken muscle and liver is irreversibly inactivated by urea. The purification of enolase was partial and resulted in a low yield. No enzymatic activity was detected for either recombinant and native enolases, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, suggesting that X. fastidiosa uses the Entner-Doudoroff pathway to produce pyruvate. Evidence is presented supporting the idea that the regulation of genes and the presence of isoforms with regulation patterns might make it difficult to understand the metabolism of carbohydrates in X. fastidiosa