Application of extracellular polymers on soil communities exposed to oil and nickel contamination.

The petrochemical industry is responsible for many accidental releases of pollutants in soil such as hydrocarbons and toxic metals. This co-contamination is responsible for a delay in the degradation of the organic pollution. Many successful technologies to remove these metals apply extracellular polymeric substances (EPS). In this study, we tested the application of an EPS from a Paenibacillus sp. to aid the bioremediation of soils contaminated with crude oil and nickel. We conducted a microcosm experiment to soils containing combinations of oil, nickel, and EPS. The final concentration of oil was evaluated with an infrared spectrometer. Also, we sequenced the metagenomes of the samples in an ion torrent sequencer. The application of EPS did not aid the removal of hydrocarbons with or without the presence of nickel. However, it led to a smaller decrease in the diversity indexes. EPS decreased the abundance of Actinobacteria and increased that of Proteobacteria. The EPS also decreased the connectivity among Actinobacteria in the network analysis. The results indicated that the addition of EPS had a higher effect on the community structure than nickel. Altogether, our results indicate that this approach did not aid the bioremediation of hydrocarbons likely due to its effect in the community structure that affected hydrocarbonoclastic microorganisms.

Influence of different sources of contamination on the microbiological quality of lettuce in the Teresópolis region, RJ, Brazil / Influência de diferentes fontes de contaminação na qualidade microbiológica de alface na região de Teresópolis, RJ

ABSTRACT Irrigation water and cultivated soil have been identified as possible sources of contamination in several crops. In certain vegetables that are eaten raw, such as lettuce, this contamination can lead to public health problems. Aiming to evaluate the influence of these sources on the quality of lettuce grown in the Córrego Sujo Basin, Teresópolis, RJ, an important agricultural pole whose production services the metropolitan region of Rio de Janeiro, water from different sources (spring, weir and river) was collected in this region, as well as samples of soil and lettuce irrigated with these waters, to carry out conventional microbiological analyzes (counts of total heterotrophic bacteria and thermotolerant coliforms) and molecular analyzes (PCR-DGGE). The count of fecal coliforms in lettuce suggests that there is an influence of irrigation water and the cultivated soil on the contamination of these vegetables. The grouping of bacterial communities in the different samples obtained by the PCR-DGGE technique shows that irrigation water has a greater influence on the contamination of these vegetables in relation to the soil where they are grown. These results corroborate the need to monitor water bodies used for irrigation and demonstrate that the PCR-DGGE technique is of great value for the study of microbial communities and, when associated with specific primers, can help in the detection of pathogens in food.

RESUMO A água de irrigação e o solo de cultivo têm sido apontados como possíveis fontes de contaminação em diversas culturas. Em determinadas hortaliças consumidas cruas, como a alface, essa contaminação pode causar problemas de saúde pública. Objetivando avaliar a influência dessas fontes na qualidade das alfaces cultivadas na Bacia do Córrego Sujo, Teresópolis, RJ; importante polo agrícola com produção voltada à região metropolitana do Rio de Janeiro, coletou-se nesta região águas proveniente de diferentes fontes (nascente, açude e rio); solos e alfaces irrigados com essas águas, para realização de análises microbiológicas convencionais (contagens de bactérias heterotróficas totais e coliformes termotolerantes) e moleculares (PCR-DGGE). A contagem de coliformes fecais na alface sugere que existe influência da água de irrigação e do solo na contaminação desses vegetais. O agrupamento das comunidades bacterianas nas diferentes amostras obtido pela técnica de PCR-DGGE mostra que a água de irrigação tem influência maior na contaminação dessas hortaliças em relação ao solo onde são cultivadas. Esses resultados corroboram a necessidade de monitoramento de corpos d'água utilizados para irrigação e demonstram ser a técnica do PCR-DGGE de grande valia para o estudo das comunidades microbianas e, quando associada a iniciadores específicos podem ajudar na detecção de patógenos em alimentos.

Effect of the solid state fermentation of cocoa shell on the secondary metabolites, antioxidant activity, and fatty acids.

During cocoa (Theobroma cacao L.) processing, the accumulated cocoa shell can be used for bioconversion to obtain valuable compounds. Here, we evaluate the effect of solid-state fermentation of cacao flour with Penicillium roqueforti on secondary metabolite composition, phenol, carotenoid, anthocyanin, flavonol, and fatty acids contents, and antioxidant activity. We found that the total concentrations of anthocyanins and flavonols did not change significantly after fermentation and the phenolic compound and total carotenoid concentrations were higher. The fermentation process produced an increase in saponin concentration and antioxidant activity, as well as significant changes in the levels of oleic, linoleic, gamma-linolenic, and saturated fatty acids. Based on our findings, we propose that the reuse of food residues through solid state fermentation is viable and useful.

The influence of nickel on the bioremediation of multi-component contaminated tropical soil: microcosm and batch bioreactor studies.

Large petrochemical discharges are responsible for organic and inorganic pollutants in the environment. The purpose of this study was to evaluate the influence of nickel, one of the most abundant inorganic element in crude oil and the main component of hydrogen catalysts for oil refining, on the microbial community structure in artificially petroleum-contaminated microcosms and in solid phase bioreactor studies. In the presence of metals, the oil biodegradation in microcosms was significantly delayed during the first 7 days of operation. Also, increasing amounts of moisture generated a positive influence on the biodegradation processes. The oil concentration, exhibiting the most negative influence at the end of the treatment period. Molecular fingerprinting analyses (denaturing gradient gel electrophoresis--DGGE) indicated that the inclusion of nickel into the contaminated soil promoted direct changes to the microbial community structure. By the end of the experiments, the results of the total petroleum hydrocarbons removal in the bioreactor and the microcosm were similar, but reductions in the treatment times were observed with the bioreactor experiments. An analysis of the microbial community structure by DGGE using various markers showed distinct behaviors between two treatments containing high nickel concentrations. The main conclusion of this study was that Nickel promotes a significant delay in oil biodegradation, despite having only a minor effect over the microbial community.

Development and validation of automatic HS-SPME with a gas chromatography-ion trap/mass spectrometry method for analysis of volatiles in wines.

An automated headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-ion trap/mass spectrometry (GC-IT/MS) was developed in order to quantify a large number of volatile compounds in wines such as alcohols, ester, norisoprenoids and terpenes. The procedures were optimized for SPME fiber selection, pre-incubation temperature and time, extraction temperature and time, and salt addition. A central composite experimental design was used in the optimization of the extraction conditions. The volatile compounds showed optimal extraction using a DVB/CAR/PDMS fiber, incubation of 5 ml of wine with 2g NaCl at 45 °C during 5 min, and subsequent extraction of 30 min at the same temperature. The method allowed the identification of 64 volatile compounds. Afterwards, the method was validated successfully for the most significant compounds and was applied to study the volatile composition of different white wines.

High-Yield Endoglucanase Production by Trichoderma harzianum IOC-3844 Cultivated in Pretreated Sugarcane Mill Byproduct.

The low-cost production of cellulolytic complexes presenting high action at mild conditions and well-balanced cellulase activities is one of the major bottlenecks for the economical viability of the production of cellulosic ethanol. In the present paper, the filamentous fungus Trichoderma harzianum IOC-3844 was used for the production of cellulases from a pretreated sugarcane bagasse (namely, cellulignin), by submerged fermentation. This fungal strain produced high contents of endoglucanase activity (6,358 U·L(-1)) after 72 hours of process, and further relevant ß-glucosidase and FPase activities (742 and 445 U·L(-1), resp.). The crude enzyme extract demonstrated appropriate characteristics for its application in cellulose hydrolysis, such as high thermal stability at up to 50°C, accessory xylanase activity, and absence of proteolytic activity towards azocasein. This strain showed, therefore, potential for the production of complete cellulolytic complexes aiming at the saccharification of lignocellulosic materials.

Trichoderma harzianum IOC-4038: A promising strain for the production of a cellulolytic complex with significant ß-glucosidase activity from sugarcane bagasse cellulignin.

Sugarcane bagasse is an agroindustrial residue generated in large amounts in Brazil. This biomass can be used for the production of cellulases, aiming at their use in second-generation processes for bioethanol production. Therefore, this work reports the ability of a fungal strain, Trichoderma harzianum IOC-4038, to produce cellulases on a novel material, xylan free and cellulose rich, generated from sugarcane bagasse, named partially delignified cellulignin. The extract produced by T. harzianum under submerged conditions reached 745, 97, and 559 U L(-1) of ß-glucosidase, FPase, and endoglucanase activities, respectively. The partial characterization of this enzyme complex indicated, using a dual analysis, that the optimal pH values for the biocatalysis ranged from 4.9 to 5.2 and optimal temperatures were between 47 and 54 °C, depending on the activity studied. Thermal stability analyses revealed no significant decrease in activity at 37 °C during 23 h of incubation. When compared to model strains, Aspergillus niger ATCC-16404 and Trichoderma reesei RutC30, T. harzianum fermentation was faster and its extract showed a better balanced enzyme complex, with adequate characteristics for its application in simultaneous saccharification and fermentation processes.

Bioemulsifier production by Microbacterium sp. strains isolated from mangrove and their application to remove cadmiun and zinc from hazardous industrial residue

The contamination of ecosystems with heavy metals is an important issue in current world and remediation technologies should be in according to environmental sustainability concept. Bioemulsifier are promising agents to be used in metal removal and could be effective to many applications in environmental industries. The aims of this work was screening the potential production of bioemulsifier by microorganisms isolated from an oil contaminated mangrove, and evaluate cadmium and zinc removal potential of those strains from a hazardous industrial residue. From that, bioemulsifier-producing bacteria were isolated from urban mangrove sediments. Four isolates were identified as Microbacterium sp by 16S rRNA analysis and were able to reduce up to 53.3 percent of culture medium surface tension (TS) when using glucose as carbon and energy source and 20.2 percent when sucrose was used. Suspensions containing bioemulsifier produced by Microbacterium sp. strains show to be able to remove cadmium and zinc from contaminated industrial residue, and its ability varied according carbon source. Significant differences in metal removal were observed by all strains depending on the carbon source. When glucose was used, Cd and Zn removal varied from 17 to 41 percent, and 14 to 68 percent, respectively. However, when sucrose was used it was observed only 4 to a maximum of 15 percent of Cd removal, and 4 to 17 percent of Zn removal. When the same tests were performed after ethanol precipitation, the results were different: the percentages of removal of Zn (7-27 percent) and Cd (14-32 percent) were higher from sucrose cultures. This is the first report of heavy metals removal by bioemulsifier from Microbacterium sp.

Bioemulsifier production byMicrobacterium SP. strains isolated from mangrove and their application to remove cadmiun and zinc from hazardous industrial residue.

The contamination of ecosystems with heavy metals is an important issue in current world and remediation technologies should be in according to environmental sustainability concept. Bioemulsifier are promising agents to be used in metal removal and could be effective to many applications in environmental industries. The aims of this work was screening the potential production of bioemulsifier by microorganisms isolated from an oil contaminated mangrove, and evaluate cadmium and zinc removal potential of those strains from a hazardous industrial residue. From that, bioemulsifier-producing bacteria were isolated from urban mangrove sediments. Four isolates were identified as Microbacterium sp by 16S rRNA analysis and were able to reduce up to 53.3% of culture medium surface tension (TS) when using glucose as carbon and energy source and 20.2% when sucrose was used. Suspensions containing bioemulsifier produced by Microbacterium sp. strains show to be able to remove cadmium and zinc from contaminated industrial residue, and its ability varied according carbon source. Significant differences in metal removal were observed by all strains depending on the carbon source. When glucose was used, Cd and Zn removal varied from 17 to 41%, and 14 to 68%, respectively. However, when sucrose was used it was observed only 4 to a maximum of 15% of Cd removal, and 4 to 17% of Zn removal. When the same tests were performed after ethanol precipitation, the results were different: the percentages of removal of Zn (7-27%) and Cd (14-32%) were higher from sucrose cultures. This is the first report of heavy metals removal by bioemulsifier from Microbacterium sp.

Cellulases from Penicillium funiculosum: production, properties and application to cellulose hydrolysis.

The objective of this work is to investigate the utilization of two abundant agricultural residues in Brazil for the production and application of cellulolytic enzymes. Different materials obtained after pretreatment of sugarcane bagasse, as well as pure synthetic substrates, were considered for cellulase production by Penicillium funiculosum. The best results for FPase (354 U L(-1)) and beta-glucosidase (1,835 U L(-1)) production were observed when sugarcane bagasse partially delignified cellulignin (PDC) was used. The crude extract obtained from PDC fermentation was then partially characterized. Optimal temperatures for cellulase action ranged from 52 to 58 degrees C and pH values of around 4.9 contributed to maximum enzyme activity. At 37 degrees C, the cellulases were highly stable, losing less than 15% of their initial activity after 23 h of incubation. There was no detection of proteases in the P. funiculosum extract, but other hydrolases, such as endoxylanases, were identified (147 U L(-1)). Finally, when compared to commercial preparations, the cellulolytic complex from P. funiculosum showed more well-balanced amounts of beta-glucosidase, endo- and exoglucanase, resulting in the desired performance in the presence of a lignocellulosic material. Cellulases from this filamentous fungus had a higher glucose production rate (470 mg L(-1) h(-1)) when incubated with corn cob than with Celluclast, GC 220 and Spezyme (312, 454 and 400 mg L(-1) h(-1), respectively).

Application of a bacterial extracellular polymeric substance in heavy metal adsorption in a co-contaminated aqueous system / Aplicação de uma substância polimerica extracelular bacteriana na absorção de metais pesados em um sistema aquoso co-contaminado

The application of a bacterial extracellular polymeric substance (EPS) in the bioremediation of heavy metals (Cd, Zn and Cu) by a microbial consortium in a hydrocarbon co-contaminated aqueous system was studied. At the low concentrations used in this work (1.00 ppm of each metal), it was not observed an inhibitory effect on the cellular growing. In the other hand, the application of the EPS lead to a lower concentration of the free heavy metals in solution, once a great part of them is adsorbed in the polymeric matrix (87.12 percent of Cd; 19.82 percent of Zn; and 37.64 percent of Cu), when compared to what is adsorbed or internalized by biomass (5.35 percent of Cd; 47.35 percent of Zn; and 24.93 percent of Cu). It was noted an increase of 24 percent in the consumption of ethylbenzene, among the gasoline components that were quantified, in the small interval of time evaluated (30 hours). Our results suggest that, if the experiments were conducted in a larger interval of time, it would possibly be noted a higher effect in the degradation of gasoline compounds. Still, considering the low concentrations that were evaluated, it is possible that a real system could be bioremediated by natural attenuation process, demonstrated by the low effect of those levels of contaminants and co-contaminants over the naturally present microbial consortium.

A aplicação de uma substância polimérica extracelular (EPS) bacteriana na biorremediação de metais pesados (Cd, Zn e Cu) por um consórcio microbiano em um sistema aquoso co-contaminado com hidrocarbonetos foi estudada. Nas baixas concentrações usadas neste trabalho (1,00 ppm de cada metal), não foi observado um efeito inibitório no crescimento celular. Por outro lado, a aplicação da EPS bacteriana levou a uma menor concentração de metais livres em solução, uma vez que grande parte destes fica adsorvido na matriz polimérica (87,12 por cento de Cd; 19,82 por cento de Zn; e 37,64 por cento de Cu) quando comparado ao que é adsorvido ou interiorizado pela biomassa (5,35 por cento de Cd; 47,35 por cento de Zn; e 24,93 por cento de Cu). No pequeno intervalo de tempo avaliado (30 horas) e na baixa concentração de gasolina utilizada (0,1 por cento (v/v)), foi percebido um aumento de 24 por cento no consumo de etilbenzeno, entre os componentes da gasolina que foram quantificados. Nossos resultados sugerem que, se os experimentos fossem conduzidos em um intervalo de tempo maior, possivelmente poderia ter sido observado um maior efeito na degradação dos componentes da gasolina. Ainda, considerando as baixas concentrações avaliadas, é provável que um sistema real pudesse ser biorremediado pelo processo de atenuação natural, tendo em vista o baixo efeito desses níveis de contaminantes e co-contaminantes sobre o consórcio microbiano naturalmente presente.

Production of 6-pentyl-alfa-pyrone by trichoderma harzianum in solid-state fermentation / Produção de 6-pentil-alfa-pirona por Trichoderma harzianum em fermentação em estado sólido

Many Trichoderma species are able to produce 6-pentyl-α-pyrone (6-PP), a lactone with coconut-like aroma. In the present work, several culture parameters were studied to enhance the production of 6-PP by Trichoderma harzianum 4040 in solid-state fermentation. Green coir powder added to a nutrient solution was used as support material for fermentation. A Plackett-Burman screening technique was applied, followed by a fractionary factorial design. The best culture conditions within the experimental domain studied were (100 g support)-1: sucrose, 3 g; NaNO3, 0.24 g; (NH4)2SO4, 0.18 g; KH2PO4, 0.1 g; inoculum concentration, 2.2 x 10(6) spores; moisture level, 55 percent. The temperature established was 28ºC. The fermentation under the selected conditions led to a 6-PP production six times higher (5.0 mg/g dry matter) than the initial one (0.8 mg/g dry matter) after seven days of cultivation.

Muitas espécies do gênero Trichoderma são capazes de produzir a substância 6-pentil-α-pirona (6-PP), uma lactona com aroma característico de coco. No presente trabalho, vários parâmetros de cultura foram estudados para aumentar a produção de 6-PP por Trichoderma harzianum 4040 em fermentação em estado sólido. Pó da casca de coco verde adicionado à uma solução nutriente foi usado como material de suporte para a fermentação. Um planejamento experimental de varredura segundo a técnica de Plackett-Burman foi aplicado, seguido de um planejamento fatorial fracionário. No domínio experimental estudado, as melhores condições de cultura foram (100 g suporte)-1: sacarose, 3 g; NaNO3, 0,24 g; (NH4)2SO4, 0,18 g; KH2PO4, 0,1 g; produção do inóculo, 2,2 x 10(6) esporos; umidade, 55 por cento. A temperatura estabelecida foi de 28ºC. Esse estudo conduziu à concentração de 6-PP seis vezes maior (5,0 mg/g de matéria seca) do que a inicial (0,8 mg/g de matéria seca) após sete dias de cultivo.

Production of 6-pentyl-α-pyrone by trichoderma harzianum in solid-state fermentation.

Many Trichoderma species are able to produce 6-pentyl-α-pyrone (6-PP), a lactone with coconut-like aroma. In the present work, several culture parameters were studied to enhance the production of 6-PP by Trichoderma harzianum 4040 in solid-state fermentation. Green coir powder added to a nutrient solution was used as support material for fermentation. A Plackett-Burman screening technique was applied, followed by a fractionary factorial design. The best culture conditions within the experimental domain studied were (100 g support)(-1): sucrose, 3 g; NaNO3, 0.24 g; (NH4)2SO4, 0.18 g; KH2PO4, 0.1 g; inoculum concentration, 2.2 × 10(6) spores; moisture level, 55%. The temperature established was 28°C. The fermentation under the selected conditions led to a 6-PP production six times higher (5.0 mg/g dry matter) than the initial one (0.8 mg/g dry matter) after seven days of cultivation.

Application of a bacterial extracellular polymeric substance in heavy metal adsorption in a co-contaminated aqueous system.

The application of a bacterial extracellular polymeric substance (EPS) in the bioremediation of heavy metals (Cd, Zn and Cu) by a microbial consortium in a hydrocarbon co-contaminated aqueous system was studied. At the low concentrations used in this work (1.00 ppm of each metal), it was not observed an inhibitory effect on the cellular growing. In the other hand, the application of the EPS lead to a lower concentration of the free heavy metals in solution, once a great part of them is adsorbed in the polymeric matrix (87.12% of Cd; 19.82% of Zn; and 37.64% of Cu), when compared to what is adsorbed or internalized by biomass (5.35% of Cd; 47.35% of Zn; and 24.93% of Cu). It was noted an increase of 24% in the consumption of ethylbenzene, among the gasoline components that were quantified, in the small interval of time evaluated (30 hours). Our results suggest that, if the experiments were conducted in a larger interval of time, it would possibly be noted a higher effect in the degradation of gasoline compounds. Still, considering the low concentrations that were evaluated, it is possible that a real system could be bioremediated by natural attenuation process, demonstrated by the low effect of those levels of contaminants and co-contaminants over the naturally present microbial consortium.

Tanase: conceitos, produção e aplicação / Tannase: concepts, production and application

O objetivo deste trabalho foi efetuar revisão do conhecimento sobre tanase, apontando necessidades e perspectivas de utilização, em especial para a área de alimentos. Foram enfatizados tópicos como importância fisiológica da tanase e seu mecanismo de ação, sua produção por microorganismos, os métodos analíticos para sua determinação, a sua produção por fermentãção, a regulação da biosíntese, suas características e propriedades, a imobilização e suas aplicações. A principal aplicação dessa enzima encontra-se na produção de chás instantâneos, porém outros campos potenciais podem ser vislumbrados como sua utilização para elaboração de rações animais com maior valor nutricional, produção de compostos antioxidantes e produçaõ de ácido gálico

Nitrification of effluents: microbiological characterization

This work is concerned with the microbiological monitoring of nitrification processes carried out in two distinct continuous reactors: submerse fixed-bed and airlift fluidized bad reactor. In the first case the system was operated using a synthetic effluent with variable salinity while in the second an actual petroleum refinery effluent was tested. On the basis of analys is for ammonia, nitrite and nitrate and nitrate at the inflow and outflow streams, system performance was evaluate. By inoculating appropriate solid medium, biotypes were isolated and colonies morphology was characterized. In general, colony of ammonia oxidizin bacteria (group 1) were round, initially transparent later, but becoming whitsh or phosphorescent. It took 36 to 48 hours for those colony to develop. The nitrite oxidizing bacteria (group 2) were generally white or redish-brown with colonies, developing time between 24 and 36 hours. In spite of the existence of various types of colonies, lamina observation under microscope revealed mainly Gram (-) bacilli with varied size. Typical width X length for individual bacterium cells were as follows: 1 X 1,5mm for group 1, normally arranged in chains, and 0,5 X 1,0mm for group 2, generally shaped as small aggregates.

Gasoline biodegradation in different soil microcosms

The objective of this study was to evaluate gasoline bidegradation in batch soil microcosms. Microorganisms able to grow in the presence of gasoline were isolated from soil. Several treatment systems were performed using both isolated strains and Pseudomonas putida obtained from a culture collection. The treatment system using only autochthonous microflora (system 1) presented an average value of degradation of 50(per cent). The association of Pseudomonas putida, Burkholderia cepacia, Pseudomonas alcaligenes and the native soil microflora (system 13) presented significant percentage of removal of n-undecane (88.7), n-dodecane(61.3) and n-tridecane(66.7). According to these results, system 1 and 13 revealed considerable potential for application in biorremediation treatments.

Accumulation of cadmium from moderately concentrated cadmium solutions by chlorella and scenedesmus strains

A acumulaçäo de Cadmio a partir de soluçöes contaminadas com o metal foi estudada através do emprego de 3 diferentes microalgas. Dentre as espécies testadas a Chlorella homosphaera mostrou maior poder de acumulaçäo comparada com células de Scenedesmus quadricauda e Chlorella homosphaera e Scenedesmus quadricauda foram consideravelmente afetadas pela presen