Benzoate degradation by Rhodococcus opacus 1CP after dormancy: Characterization of dioxygenases involved in the process

The process of benzoate degradation by strain Rhodococcus opacus 1CP after a five-year dormancy was investigated and its peculiarities were revealed. The strain was shown to be capable of growth on benzoate at a concentration of up to 10 g L^?1. The substrate specificity of benzoate dioxygenase (BDO) during the culture growth on a medium with a low (200–250 mg L^?1) and high (4 g L^?1) concentration of benzoate was assessed. BDO of R. opacus 1CP was shown to be an extremely narrow specificity enzyme. Out of 31 substituted benzoates, only with one, 3-chlorobenzoate, its activity was higher than 9% of that of benzoate. Two dioxygenases, catechol 1,2-dioxygenase (Cat 1,2-DO) and protocatechuate 3,4-dioxygenase (PCA 3,4-DO), were identified in a cell-free extract, purified and characterized. The substrate specificity of Cat 1,2-DO isolated from cells of strain 1CP after the dormancy was found to differ significantly from that of Cat 1,2-DO isolated earlier from cells of this strain grown on benzoate. By its substrate specificity, the described Cat 1,2-DO was close to the Cat 1,2-DO from strain 1CP grown on 4-methylbenzoate. Neither activity nor inhibition by protocatechuate was observed during the reaction of Cat 1,2-DO with catechol, and catechol had no inhibitory effect on the reaction of PCA 3,4-DO with protocatechuate.     

Aerobic degradation of di- and trichlorobenzenes by two bacteria isolated from polluted tropical soils

Two polychlorinated biphenyl (PCBs)-degrading bacteria were isolated by traditional enrichment technique from electrical transformer fluid (Askarel)-contaminated soils in Lagos, Nigeria. They were classified and identified as Enterobacter sp. SA-2 and Pseudomonas sp. SA-6 on the basis of 16S rRNA gene analysis, in addition to standard cultural and biochemical techniques. The strains were able to grow extensively on dichloro- and trichlorobenzenes. Although they failed to grow on tetrachlorobenzenes, monochloro- and dichlorobenzoic acids, they were able to utilize all monochlorobiphenyls, and some dichlorobiphenyls as sole sources of carbon and energy. The effect of incubation with axenic cultures on the degradation of 0.9 mM 1,4-dichlorobenzene, 0.44 mM 1,2,3- and 0.43 mM 1,3,5-trichlorobenzene in mineral salts medium was studied. Approximately, 80-90% of these xenobiotics were degraded in 200 h, concomitant with cell increase of up to three orders of magnitude, while generation times ranged significantly (P<0.05) from 17-32 h. Catechol 1,2-dioxygenase and catechol 2,3-dioxygenase activities were detected in crude cell-free extracts of cultures pre-grown with benzoate, with the latter enzyme exhibiting a slightly higher activity (0.15-0.17 micromolmin(-1) mg of protein(-1)) with catechol, suggesting that the meta-cleavage pathway is the most readily available catabolic route in the SA strains. The wider substrate specificity of these tropical isolates may help in assessing natural detoxification processes and in designing bioremediation and bioaugmentation methods.     

The effect of cyclic aerobic-anoxic conditions on biodegradation of benzoate.

The response of a mixed microbial culture to cyclic aerobic and anoxic (denitrifying) conditions was studied in a chemostat with a 48-hour hydraulic residence time receiving a feed containing benzoate and pyruvate. When the cyclic conditions were 3-hour aerobic and 9-hour anoxic, the bacteria-degraded benzoate aerobically via the catechol 2,3-dioxygenase (C23DO) pathway. The quantity of C23DO remained constant throughout the anoxic period but decreased during the initial portion of the aerobic period before returning to the level present in the anoxic period. Anoxic biodegradation of benzoate was via benzoyl-CoA reductase, which remained constant regardless of the redox condition. The aerobic benzoate uptake capability (AeBUC) of the culture increased during the aerobic period but decreased during the anoxic period. The anoxic benzoate uptake capability (AnBUC) exhibited the opposite response. When the cycle was 6-hour aerobic and 6-hour anoxic, aerobic biodegradation of benzoate proceeded via the protocatechuate 4,5-dioxygenase (P45DO) pathway. The P45DO activity decreased early in the aerobic period, but then increased to the level present during the anoxic period. The level of benzoyl-CoA reductase was constant throughout the cycle. Furthermore, AeBUC and AnBUC responded in much the same way as in the 3/9-hour chemostat. During a 9-hour aerobic and 3-hour anoxic cycle, the culture synthesized both P45DO and C23DO, with the former having significantly higher activity. Unlike the other two cycles, AeBUC changed little during the aerobic period, although AnBUC decreased. The culture was well-adapted to the cyclic conditions as evidenced by the lack of accumulation of either substrate during any cycle tested. This suggests that cyclic aerobic-anoxic processes can be used in industrial wastewater-treatment facilities receiving significant quantities of simple aromatic compounds like benzoate. However, the results showed that the kinetics of benzoate degradation were different under aerobic and anoxic conditions, a situation that must be considered when modeling cyclic bioreactors receiving aromatic compounds.     

Biodegradation and removal of 3,4-dichloroaniline by Chlorella pyrenoidosa based on liquid chromatography-electrospray ionization-mass spectrometry

3,4-Dichloroaniline (3,4-DCA), widely used in the synthesis of dyes, textile and herbicides, is toxic to living organisms. The purpose of this study was to investigate the capability of green algae in degrading and removing 3,4-DCA in water. An environmentally ubiquitous green alga Chlorella pyrenoidosa was isolated from fresh aquatic environment. Then unicellular alga was incubated with 3,4-DCA at a concentration of 4.6 μg/?mL in water. The residual concentration of 3,4-DCA in the medium and the metabolites were analyzed. A removal percentage of 78.4 % was obtained over a 7-day period. Two major metabolites with less toxicity were identified as 3,4-dichloroformanilide and 3,4-dichloroacetanilide from the liquid chromatography-electrospray ionization-mass spectrometry analysis. The application of microalga C. pyrenoidosa may have potential for removing the environmental pollutant in aquatic environment.     

Sorption of 3,4-dichloroaniline on four contrasting Greek agricultural soils and the effect of liming

Sorption of 3,4-dichloroaniline (3,4-DCA) on four typical Greek agricultural soils, with distinct texture, organic matter content and cation exchange capacities, was compared by using sorption isotherms and the parameters calculated from the fitted Freundlich equations. The sorption process of 3,4-DCA to the soil was completed within 48–72 h. The 3,4-DCA sorption on all soils was well described by the Freundlich equation and all sorption isotherms were of the L-type. The sandy clay loam soil with the highest organic matter content and a slightly acidic pH was the most sorptive, whereas the two other soil types, a high organic matter and neutral pH clay and a low organic matter and acidic loam, had an intermediate sorption capacity. A typical calcareous soil with low organic matter had the lowest sorption capacity which was only slightly higher than that of river sand. The 3,4-DCA sorption correlated best to soil organic matter content and not to clay content or cation exchange capacity, indicating the primary role of organic matter. The distribution coefficient (K _d) decreased with increasing initial 3,4-DCA concentration and the reduction was most pronounced with the highly sorptive sandy clay loam soil, suggesting that the available sorption sites of the soils are not unlimited. Liming of the two acidic soils (the sandy clay loam and the loam) raised their pH (from 6.2 and 5.3, respectively) to 7.8 and reduced their sorption capacity by about 50 %, indicating that soil pH may be the second in importance factor (after organic matter) determining 3,4-DCA sorption.     

Effects of dissolved organic carbon on sorption of 3,4-dichloroaniline and 4-bromoaniline in a calcareous soil

To evaluate the effects of dissolved organic carbon on sorption of 3,4-dichloroaniline (3,4-DCA) and 4-bromoaniline (4-BA) on soils, batch sorption experiments were carried out. The soil used was a typical calcareous soil from south-eastern Spain. Two different types of dissolved organic carbon were used, that is, dissolved organic carbon extracts from a commercial peat (DOC-PE) and high-purity tannic acid (DOC-TA). The experiments were carried out in a 0.01 M CaCl2 aqueous medium at 25 degrees C. The results obtained from the sorption experiments show that the presence of both DOC-PE and DOC-TA, over a concentration range of 15-100 mg l-1, produced in all cases, an increase in the amount of 3,4-DCA and 4-BA adsorbed on the soil studied.     

Sorption of 3,4-dichloroaniline on four contrasting Greek agricultural soils and the effect of liming

Sorption of 3,4-dichloroaniline (3,4-DCA) on four typical Greek agricultural soils, with distinct texture, organic matter content and cation exchange capacities, was compared by using sorption isotherms and the parameters calculated from the fitted Freundlich equations. The sorption process of 3,4-DCA to the soil was completed within 48-72 h. The 3,4-DCA sorption on all soils was well described by the Freundlich equation and all sorption isotherms were of the L-type. The sandy clay loam soil with the highest organic matter content and a slightly acidic pH was the most sorptive, whereas the two other soil types, a high organic matter and neutral pH clay and a low organic matter and acidic loam, had an intermediate sorption capacity. A typical calcareous soil with low organic matter had the lowest sorption capacity which was only slightly higher than that of river sand. The 3,4-DCA sorption correlated best to soil organic matter content and not to clay content or cation exchange capacity, indicating the primary role of organic matter. The distribution coefficient (K(d)) decreased with increasing initial 3,4-DCA concentration and the reduction was most pronounced with the highly sorptive sandy clay loam soil, suggesting that the available sorption sites of the soils are not unlimited. Liming of the two acidic soils (the sandy clay loam and the loam) raised their pH (from 6.2 and 5.3, respectively) to 7.8 and reduced their sorption capacity by about 50 %, indicating that soil pH may be the second in importance factor (after organic matter) determining 3,4-DCA sorption.     

两相厌氧流化床中优势菌种降解硝基苯废水的特性

构建了从强化传质与优势菌相结合的两相厌氧流化床生物降解体系,考察了水力停留时间(HRT)与上流速度2种水力特征以及共基质、pH、进水浓度等主要过程因素对优势菌种降解硝基苯的影响.结果显示,反应器在HRT为36h、上流速度为4 m/h时获得较好的处理效果;菌种需要pH 7.5的条件下以葡萄糖为共基质降解硝基苯,且两者的最佳质量比约为6;当进水硝基苯浓度为50～345 mg/L时,对硝基苯平均降解率和降解速率分别达到91.1%和120.9 mg/(L·d),且可耐受2.5倍以内的浓度负荷冲击.由此表明良好的反应器水力条件及优势菌种的结合可使高毒性的硝基苯在厌氧条件下有效地降解.     

Growth rate effects, responses of antioxidant enzymes and metabolic fate of the herbicide Propanil in the aquatic plant Lemna minor

Propanil (3,4-dichloropropionanilide) is a selective contact pesticide, recommended for post-emergence use in rice. This herbicide may end up in surface waters and present potential risk for aquatic vascular plants. Therefore, its toxicity was evaluated on Lemna minor L., an aquatic plant regularly used for toxicological studies, during time- and concentration-dependent exposure. Toxicity assessments were based on inhibition of growth of L. minor cultures after 24 days. The obtained results showed that the growth of Lemna was affected by the herbicide. The responses of the guaiacol peroxidase (G-POD) and glutathione S-transferase (GST) involved in the xenobiotic metabolism and antioxidative system were also investigated following Propanil exposure. Our results showed that Propanil has not induced enzymatic antioxidative defenses of L. minor. Both 3,4-dichloroaniline (3,4-DCA) and 3,4-dichloroacetanilide are the major metabolites in this plant. On the contrary, only 3,4-DCA was found in culture media after 4 days. Probably, the enzymatic hydrolysis by acyl acylamidase and the acetylation by acetyl-CoA are the major pathways for these transformation products, respectively. The results of this study showed that the selected aquatic plant has the potential to accumulate and metabolize rice herbicide, like Propanil. Based on these toxicity data this herbicide should impair the establishment of non-target aquatic plants.     

Comparison of degradation studies in a biosimulator by continuous cultivation of Pseudomonas and indigenous microorganisms with varied dissolved oxygen concentrations

The aim of this study was to compare the potential of a pure bacterial culture of Pseudomonas with the potential of an indigenous microorganism for malathion degradation by continuous cultivation using two different sets of conditions (with-culture: Pseudomonas, and without-culture: indigenous microorganisms) at varied dissolved oxygen (DO) concentrations (4.0 and 8.0 mg/L) in a biosimulator. The samples were analysed for pH, DO and COD (unfiltered). Gas chromatography (GC) revealed that almost 90% of malathion removal was achieved within ten hours of treatment in both studies of with-culture using 4.0 and 8.0 mg/L DO, whereas COD was considerably reduced during the treatment process and the removal efficiency was found to be above 90% in both conditions. The removal of COD was found to be in direct relation to retention time. However, the unfiltered COD and malathion removal efficiency was significantly influenced by DO. The percentage degradation of malathion using 4.0 mg/L DO, with-culture after 10 hours was 90.01%, whereas after 51 hours it was 99.03%. The without-culture studies showed percentage values which were 38.29% after 10 hours, whereas the corresponding value after 51 hours was 96.84%. Similarly, the percentage degradation of malathion using 8.0 mg/L DO, with-culture studies revealed that almost 89.40% malathion degradation was achieved after 10 hours, whereas it was 96.91% after 51 hours after inoculation of the culture. However, for the without-culture studies the value after 10 hours was 52.34%, whereas 96.86% malathion degradation was achieved after 51 hours without added culture. Although no significant variation in the pH was observed during the study the pH values increased gradually during the treatment process in all of the conditions. A critical evaluation of data presented revealed that the degradation potential of Pseudomonas was better when compared to the degradation potential of indigenous microorganisms.     

Effects of dissolved oxygen, pH, and anions on the 2,3-dichlorophenol degradation by photocatalytic reaction with anodic TiO(2) nanotube films

In this study, the highly-ordered TiO(2) nanotube (TNT) arrays on titanium sheets were prepared by an anodic oxidation method. Under UV illumination, the TNT films demonstrated the higher photocatalytic activity in terms of 2,3-dichlorophenol (2,3-DCP) degradation in aqueous solution than the conventional TiO(2) thin films prepared by a sol-gel method. The effects of dissolved oxygen (DO) and pH on the photocatalytic degradation of 2,3-DCP were investigated. The results showed that the role of DO in the 2,3-DCP degradation with the TNT film was significant. It was found that 2,3-DCP in alkaline solution was degraded and dechlorinated faster than that in acidic solution whereas dissolved organic carbon removal presented an opposite order in dependence of pH. In the meantime, some main intermediate products from 2,3-DCP degradation were identified by a (1)H NMR technique to explore a possible degradation pathway. A major intermediate, 2-chlororesorcinol, was identified from the 2,3-DCP decomposition as a new species compared to the findings in previous reports. Photocatalytic deactivation was also evaluated in the presence of individual anions (NO(3)(-), Cl(-), SO(4)(2-), and H(2)PO(4)(-)). The inhibition degree of photocatalytic degradation of 2,3-DCP caused by these anions can be ranked from high to low as SO(4)(2-)>Cl(-)>H(2)PO(4)(-)>NO(3)(-). The observed inhibition effect can be attributed to the competitive adsorption and the formation of less reactive radicals during the photocatalytic reaction.     

固定化解脂耶氏酵母间歇式处理油脂废水研究

固定化解脂耶氏酵母菌(Yarrowia lipolytica)在气升式反应器中处理色拉油浓度为2 000 mg/L、COD为3 000 mg/L的高COD油脂废水。结果表明,固定化菌对废水的最佳处理时间为6 d,可间歇式处理9批次高COD油脂废水,重复处理废水后,其对油脂的降解均在85%以上,对COD的降解在70%以上,降解后的排出水pH稳定在3.6~3.8。且处理动力学方程属于一级生化方程。结果进一步表明,固定化解脂耶氏酵母菌在气升式反应器中适于处理高COD含油脂的污水。     

微囊藻毒素-LR降解菌的筛选及降解特性研究

从上海市淀山湖表层水体中筛选分离出了1株降解微囊藻毒素-LR(MC-LR)的细菌并研究了其降解特性。根据细胞形态结构、生理生化特征及其16S rDNA基因序列分析,鉴定分离菌株DHU-28(GenBank序列登录号为HM047512)属嗜麦芽寡养单胞菌(Stenotrophomonas maltophilia)。微囊藻毒素降解实验结果表明,该菌株能在以MC-LR为唯一碳源、氮源的无机盐培养基中生长,6 d内可将初始质量浓度为15 mg/L的MC-LR降解为8.12 mg/L,降解效率达到45.9%。菌株DHU-28的最适生长温度是30℃,最适生长pH为7.0。酵母粉、蛋白胨、葡萄糖等营养物质可以明显促进菌株对MC-LR的降解效率,尤其是加入50 mg/L酵母粉后,6 d降解率达到63.2%。     

曝气量对悬浮系统单级自养脱氮性能的影响

通过连续实验和间歇实验研究了不同曝气量对SBR系统自养脱氮性能的影响。连续实验表明,在进水氨氮浓度为155～185 mg/L时,曝气量分别为20、28、36和44 L/h时,TN去除率分别为80%、82%、80%和77%;增大和减小曝气量均会降低系统的脱氮效率。间歇实验表明,随着曝气量的增加,氨氮的降解速率有所升高,20、28、36和44 L/h曝气条件下氨氮的降解速率分别为7.23、7.25、7.86和7.95 mg/(g MLVSS.h);在降解的过程中DO浓度一直维持在较低的水平(<0.5 mg/L),pH值则呈先升高后降低的趋势;氨氮降解结束时,pH值和DO浓度同时升高。结果表明,改变曝气量会影响单级自养脱氮反应的进程,但对降解过程DO浓度值变化不大;DO浓度和pH值变化对氨降解结束具有指示作用。     

Metabolic pathways of polychlorinated biphenyls degradation by Pseudomonas sp. 2

Polychlorinated biphenyls (PCBs) included with the commercial mixture Delor 103 were degraded by immobilized cells of aerobic bacterial strain Pseudomonas sp. 2. The ability of the strain to metabolise selected tri- and tetrachlorobiphenyls, and the site of primary attack of the biphenyl skeleton were investigated. It was observed that the amount of residual PCBs was 1-48% of the original PCBs after three weeks of incubation. Identified metabolites indicate that the used bacterial strain attacks the biphenyl skeleton at the 2,3- and 3,4-positions, and it is also able to dehalogenate PCBs. Metabolic pathways of degradation of individual congeners were proposed. Transformation of 2,4- and 2,5-dichlorobenzoic acids by Pseudomonas sp. 2 was also observed.     

Mutant AFM 2 of Alcaligenes faecalis for phenol biodegradation using He-Ne laser irradiation

He-Ne laser technology was utilized in this study to investigate the response of Alcaligenes faecalis to laser stimulation. The irradiation experiments were conducted by the adjustment of the output power from 5 to 25 mW and the exposure time from 5 to 25 min. The results showed that the survival rate changed regularly with the variety of irradiation dose, and high positive mutation frequency was determined by both the energy density and the output power. The mutant strain AFM 2 was obtained. Phenol biodegradation assay demonstrated that AFM 2 possessed a more prominent phenol-degrading potential than its parent strain, which presumably attributed to the improvements of phenol hydroxylase and catechol 1,2-dioxygenase activities. The phenol of 2000 mgl(-1) was completely degraded by AFM 2 within 85.5h at 30 degrees C. In addition, the cell growth and phenol degradation kinetics of the mutant strain AFM 2 and its parent strain in batch cultures were also investigated at the wide initial phenol concentration ranging from 0 to 2000 mgl(-1) by Haldane model. The results of these experiments further demonstrated that the mutant strain AFM 2 possessed a higher capacity to resist phenol.     

共代谢基质对苯酚降解菌XTT-3降酚作用的影响

采用驯化的方法从活性污泥中分离到一株苯酚降解菌XTT-3,经16SrDNA鉴定为Sphingobiumsp.。对该菌株进行碳饥饿处理,发现其降解苯酚的能力受到抑制。以只含苯酚的M9培养基为参照,添加0.2g/L酵母膏作为共代谢基质,对XTT-3菌株降解苯酚有较明显的促进作用,36h后苯酚降解率为68%。在含0.2g/L酵母膏的M9培养基中,同时添加20mg/L邻苯二酚,XTT-3降解苯酚作用显著增加,24h后苯酚降解率达75%,苯酚降解速度达0.261mg/min。     

Bioremediation of environmental endocrine disruptor di-n-butyl phthalate ester by Rhodococcus ruber

In this study DBP-degradation strain CQ0301 was isolated from rubbish landfill soil. According to the biophysical, biochemical characteristics and analysis of 16S rRNA, the strain was identified as Rhodococcus ruber. Three new protein bands could be fractioned after DBP-inducing, which were suspected to participate the process of DBP-degrading. Catechol was suspected to be an intermediate product of DBP and cleaving the benzene ring was catalyzed by catechol 1,2-dioxygenase, because a highly activity of catechol 1,2-dioxygenase could be detected after DBP-inducing. The results of this study also showed the optimal pH value, optimal temperature which influenced the degradation rate in soil: pH 7.0-8.0, 30-35 degrees C. Kinetics of degradation reaction had been performed at different initial concentration and different time. Analyzed with SPSS10.0 software, the DBP degradation can be described as the same exponential model when the initial DBP concentration was lower than 50 mg/kg. The kinetics equation was lnC=-0.1332t + A, with the degradation half-life of DBP in soil (5.20 d). Inoculating CQ0301 could relieve DBP content in plant. We also found that adding nutrient materials into soil was useful for decreasing the DBP content in plant. In summary, we isolated a bacterium capable of degrading DBP and decreasing DBP content in plant. We also explored the mechanism of biodegradation and characterized the environmental factors influencing the degradation process in contaminated soil. Based on this work, we hope that these findings can provide some information for applying of bioremediation of DBP contaminated soil.     

1,1-二氯乙烯降解菌的分离鉴定及降解特性

从好氧活性污泥中分离得到一株能以1,1-二氯乙烯(1,1-DCE)作为惟一碳源和能源生长的革兰氏阴性菌株D-B,经鉴定属于产碱杆菌属(Alcaligenessp.)。当维持菌株D-B浓度一定时,1,1-DCE的去除率随着1,1-DCE浓度的增大先增加后降低,且降解过程主要发生在加入1,1-DCE后的3~5 h内。当1,1-DCE的初始浓度为300μg/L时去除率达到最大值85.32%。菌株D-B对1,1-DCE的降解符合Monod方程,饱和常数Ks=21.96 mg/L,1,1-DCE的最大比基质降解速率Vmax=50.76 mg/(L.h)。     

Biodegradation of dimethylphenols by bacteria with different ring-cleavage pathways of phenolic compounds

The biodegradation of 3,4, 2,4, 2,3, 2,6 and 3,5-di-methylphenol in combination with phenol andp-cresol by axenic and mixed cultures of bacteria was investigated. The strains, which degrade phenol andp-cresol through different catabolic pathways, were isolated from river water continuously polluted with phenolic compounds of leachate of oil shale semicoke ash heaps. The proper research of degradation of 2,4 and 3,4-di-methylphenol in multinutrient environments was performed. The degradation of phenolic compounds from mixtures indicated a flux of substrates into different catabolic pathways. Catechol 2,3-dioxygenase activity was induced by dimethylphenols inPseudomonas mendocina PC1, wheremeta cleavage pathway was functional during the degradation ofp-cresol. In the case of strains PC18 and PC24 ofP. fluorescens, the degradation ofp-cresol occurred via the protocatechuateortho pathway and the key enzyme of this pathway,p-cresol methylhydroxylase, was also induced by dimethylphenols. 2,4 and 3,4-dimethylphenols were converted into the dead-end products 4-hydroxy-3-methylbenzoic acid and 4-hydroxy-2-methylbenzoic acid. In the degradation of 3,4-dimethylphenol, the transient accumulation of 4-hydroxy-2-methylbenzaldehyde repressed the consumption of phenol from substrate mixtures. A mixed culture of strains with different catabolic types made it possible to overcome the incompatibilities at degradation of studied substrate mixtures.