Centromere-associated topoisomerase activity in bloodstream form Trypanosoma brucei

Topoisomerase-II accumulates at centromeres during prometaphase, where it resolves the DNA catenations that represent the last link between sister chromatids. Previously, using approaches including etoposide-mediated topoisomerase-II cleavage, we mapped centromeric domains in trypanosomes, early branching eukaryotes in which chromosome segregation is poorly understood. Here, we show that in bloodstream form Trypanosoma brucei, RNAi-mediated depletion of topoisomerase-IIα, but not topoisomerase-IIβ, results in the abolition of centromere-localized activity and is lethal. Both phenotypes can be rescued by expression of the corresponding enzyme from T. cruzi. Therefore, processes which govern centromere-specific topoisomerase-II accumulation/activation have been functionally conserved within trypanosomes, despite the long evolutionary separation of these species and differences in centromeric DNA organization. The variable carboxyl terminal region of topoisomerase-II has a major role in regulating biological function. We therefore generated T. brucei lines expressing T. cruzi topoisomerase-II truncated at the carboxyl terminus and examined activity at centromeres after the RNAi-mediated depletion of the endogenous enzyme. A region necessary for nuclear localization was delineated to six residues. In other organisms, sumoylation of topoisomerase-II has been shown to be necessary for regulated chromosome segregation. Evidence that we present here suggests that sumoylation of the T. brucei enzyme is not required for centromere-specific cleavage activity

Treatment of textile wastewaters using combinations of biological and physico-chemical methods

Considerable amounts of water and chemicals are used in the textile industry, and the presence of dyes in the wastewater is a problem of environmental concern. Synthetic dyes are designed to resist light, water and chemicals to improve the quality of the textiles; however, these properties also contribute to their persistence in the environment. The existence of wastewater treatment plants in connection to the textile industries varies from region to region, and while some treatment plants are quite efficient, others are not even in use. The pollution of dye-containing wastewater has clear visible effects and some dyes are also suspected to be biologically modified into carcinogenic and toxic compounds. Common techniques used for the treatment of dye wastewater are physical or chemical methods, such as precipitation, membrane filtration or adsorption in combination with aerobic biological treatment. Even though these techniques can be efficient, they can also be quite expensive and are based on transfer of the pollutant from one phase to another, thereby creating secondary waste. Moreover, the majority of synthetic dyes are recalcitrant to conventional aerobic biodegradation. With the purpose of finding a better alternative for the treatment of textile wastewaters, a number of methods were evaluated using azo dyes collected from the Indian and Bolivian textile industries. The studied treatment alternatives were both biological and physico-chemical methods selected on the basis to provide complete dye degradation and not only phase removal. Since textile effluents can be very diverse depending on the concentration of the dyes and additional chemicals, the focus was also to evaluate the robustness of the methods. To confirm the recalcitrance of the dyes under conventional aerobic conditions, studies using activated sludge collected from a municipal wastewater treatment plant in Lund, Sweden, were conducted. The results showed that the dyes were not degradable under aerobic conditions. The first alternative that was evaluated was degradation by white rot fungi. These fungi grow on trees and degrade lignin through the use of extracellular enzymes. Due to the unspecificity of these enzymes, it has been shown that they also can be used to degrade a wide range of recalcitrant pollutants. The process was evaluated using various fungal species as well as extracted enzymes. Even though high decolourisation efficiencies were obtained, the dyes were not fully degraded. Moreover, it was difficult to run a stable process when using the fungi in non-sterile reactors. Secondly, we evaluated a combined anaerobic-aerobic process using sludge from a municipal wastewater treatment plant, from a textile industry and from a chemical industry. Even though this process was both robust and efficient in decolourising the dyes, all intermediates were not degraded. Thereafter two physico-chemical methods, photocatalysis and photo-Fenton, were evaluated. Both these processes are based on the production of highly reactive hydroxyl radicals, which degrade the pollutant. In the photocatalysis titanium dioxide was used as a catalyst, while hydrogen peroxide and ferrous iron were added in the photo-Fenton treatment. Almost complete dye decolourisation and degradation could be achieved using both methods. The drawbacks with these methods, however, are the high energy use and the cost of the catalyst/reagents. Combining a physico-chemical method with a biological method could take advantage of both the cost efficiency of the biological process and the efficiency of the physico-chemical process. Using combined photocatalytic-biological treatment and combined photo-Fenton-biological treatment, it was shown that complete degradation could be achieved while reducing the chemical requirements of the photo-Fenton process as well as the treatment time of the photocatalysis. Comparing the two physico-chemical methods, the photo-Fenton process appeared to be a faster and more robust alternative, requiring lower catalyst concentrations. The combined biological-photo-Fenton process can thereby be considered as a promising alternative for treatment of textile wastewaters and should be further studied on real textile wastewater using sunlight as radiation source

Treatment of synthetic textile wastewater by homogeneous and heterogeneous photo-Fenton oxidation

The efficiencies of homogeneous and heterogeneous photo-Fenton oxidation were compared for treatment of azo dye containing synthetic textile wastewater. The influence of parameters such as the presence of NaCl and starch was evaluated and optimal iron and H2O2 dosage determined. Complete decolorization of Remazol Red RR was achieved at all investigated NaCl and starch concentration. Mineralization, in terms of COD reduction, was 96% in the homogeneous and 93% in the heterogeneous reaction, but decreased with increasing NaCl and starch concentrations. The homogeneous oxidation showed the highest efficiency in treating synthetic wastewater containing Remazol Blue RR or a mixture of Remazol Red RR and Remazol Blue RR. Nevertheless, the mineralization was significantly lower than for Remazol Red RR, which shows the dependence on dye structure. Similar amounts of iron-containing sludge were produced in the two processes, while the release of iron ions was reduced by 50% when using heterogeneous photo-Fenton. Promising results were obtained when reusing the iron powder as catalyst; complete decolorization was achieved during 20 batches. (C) 2012 Elsevier B.V. All rights reserved

Comparison of anaerobic pre-treatment and aerobic post-treatment coupled to photo-Fenton oxidation for degradation of azo dyes

Photo-Fenton oxidation was used for treatment of synthetic textile wastewater as stand alone treatment, as pre-treatment before aerobic biological treatment and as post-treatment after anaerobic biological treatment. The processes were compared with regards to decolorization, chemical oxygen demand (COD) reduction and chemical consumption. When applying photo-Fenton alone for treatment of Remazol Red RR (100 mg/l), optimal conditions were 3.0 mM H2O2 and 0.25 mM Fe2+. These conditions resulted in complete decolorization and a residual COD of 2.9 mg/l. When reducing the H2O2 dose to 1 mM, residual COD was 22 mg/l. In the combined photo-Fenton/aerobic treatment complete decolorization and COD removal was achieved at 3 mM H2O2 and 0.25 mM Fe2+, while 9 mg/l of residual COD remained at the H2O2 concentration 1 mM. When applying photo-Fenton as post-treatment after the anaerobic step, the residual COD was 14 mg/l independent of the H2O2 concentration being set to 1 or 3 mM, however the Fe2+ concentration needed to be increased to 2 mM, due to complex formation with the phosphate added as a macronutrient. Phytotoxicity tests showed higher residual toxicity after the photo-Fenton treatment alone than after the combined processes. Our results thereby show that incorporation of a biological step leads to improved mineralization and reduced residual toxicity at lower H2O2 doses. (C) 2011 Elsevier B.V. All rights reserved

Immobilisation of TiO(2) for combined photocatalytic-biological azo dye degradation.

The biodegradability of the azo dye Remazol Red RR (100 mg/l) was evaluated using unadapted activated sludge and the experiment confirmed the recalcitrance of the dye. Using a combination of photocatalysis and an aerobic biological step, the biodegradability was improved significantly and complete removal of both colour and COD were achieved. Furthermore, TiO(2) was successfully immobilised on borosilicate glass slides by calcination, which facilitates reuse of the catalyst. The catalytic activity of the immobilised TiO(2) was close to that of suspended TiO(2). A reduced activity was however observed when the TiO(2) slides were used repeatedly. When comparing NaOH, calcination and UV irradiation for regeneration of the TiO(2) slides, immersion in NaOH was shown to be the most efficient method

Azo dye decolorization by a laccase/mediator system in a membrane reactor: Enzyme and mediator reusability.

This paper presents the use of a membrane-integrated reactor system with recycling of laccase and mediator for azo dye decolorization. From initial screening of different laccases and mediators, Trametes versicolor laccase and syringaldehyde provided the best system for decolorization. Decolorization yields of 98, 88, 80 and 78% were obtained for Red FN-2BL, Red BWS, Remazol Blue RR and Blue 4BL, respectively. The reaction parameters were optimized and a membrane reactor was set up for dye decolorization in batch mode with reuse of the enzyme. Between 10 and 20 batches could be run with decolorization yields from 95 to 52% depending on the dye type. To study the possibility of reusing both enzyme and mediator, the reactor was run using 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO) coupled to polyethylene glycol (PEG). Nine batches were run for the treatment of Remazol Blue RR, providing decolorization yields of 96-78%. Cost analysis of the processes showed that the costs of laccase/syringaldehyde or laccase/TEMPO were almost equal when running 20 batches, but the cost for the PEG-TEMPO was higher. However, the advantages associated with reuse of the mediator should motivate further development of the concept

Decolorization of textile dyes by Bjerkandera sp. BOL 13 using waste biomass as carbon source

BACKGROUND: Phanerochaete chrysosporium, Trametes versicolor and Bjerkandera sp BOL13 were compared for decolorization of azo dyes supplied individually or as a mixture. The dye decolorization was also evaluated during continuous treatment under non-sterile conditions using a lignocellulosic growth substrate. RESULTS: Bjerkandera sp BOL13 showed the highest dye decolorization potential. This fungus was also found to support high decolorization of Remazol Red RR at an initial pH of 4-6 and when using straw as co-substrate. The fungus was evaluated for Remazol Red RR decolorization in a continuously fed packed-bed bioreactor operated under non-sterile conditions with 3 days of hydraulic retention time. When glucose was supplied as growth-substrate, decolorization efficiencies of 65-90% were maintained for 12 days in a bioreactor packed with wooden material. The decolorization efficiency was lower when glucose was not fed to the fungus or when a plastic material was used as packing. Higher manganese peroxidase and laccase activities were also recorded when the wood packing was used. Contamination caused a drop in decolorization efficiency after 17-19 days operation. CONCLUSIONS: The potential of Bjerkandera sp BOL13 for decolorization of azo dyes under non-sterile conditions using lignocellulosic growth substrates was demonstrated. Research is needed to reduce contamination under non-sterile conditions

Degradation of a textile azo dye using biological treatment followed by photo-Fenton oxidation: Evaluation of toxicity and microbial community structure

Many commercial dye preparations are cocktails of active dyes and various by-products that are recalcitrant to biological degradation and end up in significant amounts in the effluent after the dyeing process. Conventional wastewater treatment processes are not able to degrade such compounds and detoxify the effluent, thus alternative treatments should be developed. In our work we suggest to use photo-Fenton oxidation as post-treatment after an anaerobic biofilm process, in a way to minimize the reagents needed. This process was used for treatment of synthetic textile wastewater containing the commercial azo dyestuff Remazol Red, starch and sodium chloride. The treated textile effluent had COD lower than 18 mg/l even when using initial Fenton reagents concentration as low as 1 mM ferrous ions and 10 mM hydrogen peroxide. The acute toxicity was higher in the biologically treated than in the untreated effluent. Photo-Fenton oxidation successfully reduced the toxicity and the final effluent was non-toxic to Artemia sauna and Microtox, with the exception of the effluent containing high concentration of sodium chloride, which was moderately toxic to Microtox. For the first time the presence of algae was detected in a reactor treating textile wastewater using denaturing gradient gel electrophoresis (DGGE); bacteria and fungi were also abundant. The results of this study suggest that using advanced oxidation after biological treatment is an effective way to degrade the organic compounds and remove toxicity from textile effluents. (C) 2015 Elsevier B.V. All rights reserved

Combined anaerobic-ozonation process for treatment of textile wastewater: Removal of acute toxicity and mutagenicity.

A novel set up composed of an anaerobic biofilm reactor followed by ozonation was used for treatment of artificial and real textile effluents containing azo dyes. The biological treatment efficiently removed chemical oxygen demand and color. Ozonation further reduced the organic content of the effluents and was very important for the degradation of aromatic compounds, as shown by the reduction of UV absorbance. The acute toxicity toward Vibrio fischeri and the shrimp Artemia salina increased after the biological treatment. No toxicity was detected after ozonation with the exception of the synthetic effluent containing the highest concentration, 1g/l, of the azo dye Remazol Red. Both untreated and biologically treated textile effluents were found to have mutagenic effects. The mutagenicity increased even further after 1min of ozonation. No mutagenicity was however detected in the effluents subjected to longer exposure to ozone. The results of this study suggest that the use of ozonation as short post-treatment after a biological process can be beneficial for the degradation of recalcitrant compounds and the removal of toxicity of textile wastewater. However, monitoring of toxicity and especially mutagenicity is crucial and should always be used to assess the success of a treatment strategy

MicroRNA cloning and sequencing in osteosarcoma cell lines: Differential role of miR-93

Background Studies show that abnormalities in non-coding genes can contribute to carcinogenesis; microRNA levels may modulate cancer growth and metastatic diffusion. Method MicroRNA libraries were built and sequenced from two osteosarcoma cell lines (MG-63 and 143B), which differ in proliferation and transmigration. By cloning and transfection, miR-93, expressed in both cell lines, was then investigated for its involvement in osteosarcoma progression. Results Six of the 19 miRNA identified were expressed in both cell lines with higher expression levels of miR-93 in 143B and in primary osteosarcoma cultures compared to normal osteoblasts. Interestingly, levels of miR-93 were significantly higher in metastases from osteosarcoma than in paired primary tumours. When 143B and MG-63 were transfected with miR-93, clones appeared to respond differently to microRNA overexpression. Ectopic expression of miR-93 more significantly increased cell proliferation and invasivity in 143B than in MG-63 clones. Furthermore, increased mRNA and protein levels of E2F1, one of the potential miR-93 targets, were seen in osteosarcoma cellular clones and its involvement in 143B cell proliferation was confirmed by E2F1 silencing. Conclusion Although further studies are needed to evaluate miRNA involvement in osteosarcoma progression, miR-93 overexpression seems to play an important role in osteosarcoma cell growth and invasion. © International Society for Cellular Oncology 2011