Biodegradation of azo and anthraquinone dyes in continuous systems

The purpose is to develop a complete microbiological model system for the treatment of wastewater from textile mills in developing countries. Artificial wastewater was treated by microorganisms growing on wood shavings from Norway spruce during unsterile conditions. The microorganisms were inoculated from forest residues. Mixtures of the azo dyes Reactive Black 5 and Reactive Red 2 were degraded in batch as well as continuous experiments. Reactive Red 2 mixed with the anthraquinone dye Reactive Blue 4 was also treated in the continuous system. The system consisted of three reservoirs - the first two with an anaerobic environment and the third with an aerobic. The dye concentrations were 200 mg l⁻¹ of each dye in the continuous system and the retention time was approximately 4 days and 20 h per reservoir. Samples from the process were analysed with spectrophotometer and LC/MS to monitor the degradation process. 86-90% of the colour was removed after a treatment of 4 days and 23 h in the continuous process. Two metabolites were found in the outlets of reactors one and two, but they were degraded to below the detection limit in the aerobic reactor.     

Enhanced bio-decolorization of azo dyes by co-immobilized quinone-reducing consortium and anthraquinone

In the present study, the accelerating effect of co-immobilized anthraquinone and quinone-reducing consortium was investigated in the bio-decolorization process. The anthraquinone and quinone-reducing consortium were co-immobilized by entrapment in calcium alginate. The co-immobilized beads exhibited good catalytic activity and increased the decolorization rate for many kinds of azo dyes. The reusability of the co-immobilized beads was evaluated with repeated-batch decolorization experiments. After ten repeated experiments, the decolorization rate of co-immobilized beads retained over 92.8% of their original value. Furthermore, acclimatized quinone-reducing consortium was analyzed by denaturing gradient gel electrophoresis (DGGE) and 16S rDNA gene sequencing to get the complete picture of its diversity. This study explored a great improvement of conventional treatment system and the new bio-treatment concept.     

Degradation of synthetic reactive azo dyes and treatment of textile wastewater by a fungi consortium reactor

In this paper, two microbial cultures with high decolorization efficiencies of reactive dyes were obtained and were proved to be dominant with fungi consortium in which 21 fungal strains were isolated and 8 of them showed significant decolorization effect to reactive red M-3BE. A 4.5 l continuous biofilm reactor was established using the mixed cultures to investigate the decolorization performance and the system stability under the conditions of simulated and real textile wastewater as influents. The optimal nutrient feed to this bioreactor was 0.5 g l⁻¹ glucose and 0.1 g l⁻¹ (NH₄)₂SO₄ when 30 mg l⁻¹ reactive black 5 was used as initial dye concentrations. Dye mineralization rates of 50–75% and color removal efficiencies of 70–80% were obtained at 12 h hydraulic retention time (HRT) in this case. Higher glucose concentrations in the influents could significantly improve color removal, but was not helpful for dye mineralization. Besides reactive black 5, the bioreactor could effectively decolorize reactive red M-3BE, acid red 249 and real textile wastewater with efficiency of 65%, 94% and 89%, respectively. In addition, the microbial community on the biofilm was monitored in the whole running process. The results indicated fungi as a dominant population in the decolorization system with the ratio of fungi to bacteria 6.8:1 to 51.8:1 under all the tested influent conditions. Analysis of molecular biological detection indicated that yeasts of genus Candida occupied 70% in the fungal clone library based on 26S rRNA gene sequences.     

Mutagenicity of anthraquinone and azo dyes in Ames' Salmonella typhimurium test.

23 dyes belonging to different chemical classes--anthraquinones, mono- and bis-azo compounds--were tested for their mutagenic activity on Ames strains of Salmonella typhimurium. 5 dyes induced frameshift mutations.     

Metabolites from freshwater aquatic microalgae and fungi as potential natural pesticides

Microorganisms are recognized worldwide as the major source of secondary metabolites with mega diverse structures and promissory biological activities. However, as yet many of them remain little or under-explored like the microbiota from freshwater aquatic ecosystems. In the present review, we undertook a recompilation of metabolites reported with pesticidal properties from microalgae (cyanobacteria and green algae) and fungi, specifically from freshwater aquatic habitats.     

Decolorization of triphenylmethane, azo, and anthraquinone dyes by a newly isolated Aeromonas hydrophila strain

A broad-spectrum dye-decolorizing bacterium, strain DN322, was isolated from activated sludge of a textile printing wastewater treatment plant. The strain was characterized and identified as a member of Aeromonas hydrophila based on Gram staining, morphology characters, biochemical tests, and nearly complete sequence analysis of 16S rRNA gene and the gyrase subunit beta gene (gyrB). Strain DN322 decolorized a variety of synthetic dyes, including triphenylmethane, azo, and anthraquinone dyes. For color removal, the most suitable pH and temperature were pH 5.0–10.0 and 25–37°C, respectively. Triphenylmethane dye, e.g., Crystal Violet, Basic Fuchsin, Brilliant Green, and Malachite Green (50 mg l⁻¹) were decolorized more than 90% within 10 h under aerobic culture condition and Crystal Violet could be used as sole carbon source and energy source for cell growth. The color removal of triphenylmethane dyes was due to a soluble cytosolic enzyme, and the enzyme was an NADH/NADPH-dependent oxygenase; For azo and anthraquinone dyes, e.g., Acid Amaranth, Great Red GR, Reactive Red KE-3B, and Reactive Brilliant Blue K-GR (50 mg l⁻¹) could be decolorized more than 85% within 36 h under anoxic condition. This strain may be useful for bioremediation applications.     

Decoloration of azo dyes by three whiterot fungi: influence of carbon source

Two strains of Phanerochaete chrysosporium and a local isolate of white-rot fungus, if pre-cultured in a high nitrogen medium with glucose, could decolorize two azo dyes (Amaranth and Orange G) and a heterocyclic dye (Azure B). When starch was used in the pre-cultivation medium, decoloration of Orange G occurred if the medium also contained 12mM NH₄Cl, whether or not veratric acid was present. In medium containing 1.2mM NH₄Cl and veratric acid, greater decoloration occurred with one strain of P. chrysosporium and the local isolate. In preculture medium with cellulose and 1.2mM NH₄Cl, decoloration by the local isolate was enhanced but not that by the other strains.The authors are with the Department of Microbiology, Soochow University, Shih Lin, Taipei, Taiwan     

Phylogenetic analysis of manganese-oxidizing fungi isolated from manganese-rich aquatic environments in Hokkaido, Japan

Three strains of Mn-oxidizing fungi were isolated from manganese-rich aquatic environments: sediment in a stream (Komanoyu) in Mori-machi and inflow to an artificial wetland in Kaminokuni-cho, Hokkaido, Japan. The characteristics of each strain were then established. Genetic analysis based on the ribosomal RNA (rRNA) gene was performed to clarify their classification. The sequences of the 18S rRNA and internal transcribed spacer (ITS1)-5.8S rRNA-ITS2 genes showed that all three strains are Ascomycetes. Based on its morphology, it seems probable that the KY-1 strain from Mori-machi belongs to the genus Phoma or Ampelomyces. The phylogenetic analysis indicates that this strain belongs to Phoma rather than Ampelomyces. Morphological identification of WL-1 and WL-2 strains from Kaminokuni-cho was impossible because of the lack of a sexual stage and specific organs. Phylogenetic analysis of the sequence in the ITS1-5.8S rRNA-ITS2 gene suggests that the WL-1 strain corresponds to Paraconyothyrium sporulosum and that WL-2 also belongs to the genus Paraconiothyrium. Because the ability to oxidize Mn has not been evaluated for most species of Phoma or Paraconiothyrium (Coniothyrium), further study is needed to confirm the status of these three strains.     

Biodecolorization of azo,anthraquinonic and triphenylmethane dyes by white-rot fungi and a laccase-secreting engineered strain

One laccase-secreting engineered strain and four white-rot fungi were tested for their capacity to decolorize nine dyes that could be classified as azo, anthraquinonic and triphenylmethane dyes. Trametes versicolor was the most efficient of the tested strains under these experimental conditions. Anthraquinonic dyes were decolorized more easily than the other two types. Small structural differences among the dyes could significantly affect decolorization. None of the strains showed lignin peroxidase or veratryl alcohol oxidase activity. None of the dyes were decolorized completely by laccase alone. It is likely that other phenoloxidases, such as Mn-dependent and versatile peroxidase, were also involved in decolorization of the dyes.     

Response of bioluminescent bacteria to sixteen azo dyes

Recombinant bioluminescent bacteria were used to monitor and classify the toxicity of azo dyes. Two constitutive bioluminescent bacteria,Photobacterium phosphoreum andEscherichia coli, E. coli GC2 (lac::luxCDABE), were used to detect the cellular toxicity of the azo dyes. In addition, four stress-inducible bioluminescentE. coli, DPD2794 (recA::luxCDABE), a DNA damage sensitive strain; DPD2540 (fabA::luxCDABE), a membrane damage sensitive strain; DPD2511 (katG::luxCDABE), an oxidative damage sensitive strain; and TV1061 (grpE::luxCDABE), a protein damage sensitive strain, were used to provide information about the type of toxicity caused by crystal violet, the most toxic dye of the 16 azo dyes tested. These results suggest that azo dyes result in serious cellular toxicity in bacteria, and that toxicity monitoring and classification of some azo dyes, in the field, may be possible using these recombinant bioluminescent bacteria.     

Performance of mango seed adsorbents in the adsorption of anthraquinone and azo acid dyes in single and binary aqueous solutions

In this study the husk of mango seed and two carbonaceous adsorbents prepared from it were used to study the adsorption behavior of eight acid dyes. The adsorbed amount in mmol m⁻² decayed asymptotically as the molecular volume and area increased. The interaction between the studied dyes and the mesoporous carbon was governed by the ionic species in solution and the acidic/basic groups on the surface. Less than 50% of the external surface of the microporous carbon became covered with the dyes molecules, though monolayer formation demonstrating specific interactions only with active sites on the surface and the adsorption magnitudes correlated with the shape parameter of the molecule within a particular dye group. The adsorption behavior in mixtures was determined by the molecular volume of the constituents; the greater the molecular volume difference, the greater the effect on the adsorbed amount. We also demonstrated that the raw husk of the mango seed can be used to remove up to 50% from model 50 mg l⁻¹ solutions of the studied acid dyes.     

Potentials and limitations of quantification of fungi in freshwater environments based on PLFA profiles

Aquatic fungi are increasingly recognized for their contribution to carbon cycling in aquatic ecosystems, both as saprotrophs and parasites. Their quantification in mixed communities is crucial to assess their ecological significance but remains challenging. We characterized the phospholipid-derived fatty acid (PLFA) composition of fifteen aquatic fungal isolates from Chytridiomycota (chytrids) and Dikarya. Additionally, we identified PLFA patterns of chytrids infecting phytoplankton and their zoospores. PLFA composition of zoospores was highly similar among different taxa, but were distinct from their respective sporangial life-stage. Finally, we applied a fatty acid-based Bayesian mixed model (FASTAR) and tested its potential to quantify fungi in complex mixtures with bacteria and phytoplankton using PLFA profiles. While the quantification of chytrid biomass in low quantities was rather imprecise, the model predicted the contribution of filamentous fungi and other components with fair accuracy, supporting the suitability of this approach to quantify fungal biomass in aquatic environments.     

Bacterial decolorization and degradation of azo dyes

Azo compounds constitute the largest and the most diverse group of synthetic dyes and are widely used in a number of industries such as textile, food, cosmetics and paper printing. They are generally recalcitrant to biodegradation due to their xenobiotic nature. However microorganisms, being highly versatile, have developed enzyme systems for the decolorization and mineralization of azo dyes under certain environmental conditions. Several genera of Basidomycetes have been shown to mineralize azo dyes. Reductive cleavage of azo bond, leading to the formation of aromatic amines, is the initial reaction during the bacterial metabolism of azo dyes. Anaerobic/anoxic azo dye decolorization by several mixed and pure bacterial cultures have been reported. Under these conditions, this reaction is non-specific with respect to organisms as well as dyes. Various mechanisms, which include enzymatic as well as low molecular weight redox mediators, have been proposed for this non-specific reductive cleavage. Only few aerobic bacterial strains that can utilize azo dyes as growth substrates have been isolated. These organisms generally have a narrow substrate range. Degradation of aromatic amines depends on their chemical structure and the conditions. It is now known that simple aromatic amines can be mineralized under methanogenic conditions. Sulfonated aromatic amines, on the other hand, are resistant and require specialized aerobic microbial consortia for their mineralization. This review is focused on the bacterial decolorization of azo dyes and mineralization of aromatic amines, as well as the application of these processes for the treatment of azo-dye-containing wastewaters.     

不同生境外生菌根真菌对铝胁迫的响应

以南北方不同生境下的10株外生菌根真菌为研究对象,采用液体培养的方法,研究了铝对不同菌根真菌的生物量、有机酸分泌及养分含量的影响,以期筛选出抗铝性强的优良菌株,并探讨其抗铝机理。结果表明:外生菌根真菌Sl 08抗铝性最强;Pt 715、Ld 03、Bo 11、Sl 01、Bo 15也具有不同程度的耐铝性;Sl 14、Gc 99、Cg 04抗铝性较差;Sg 11抗铝性最差。来自南方酸性森林土壤的菌株总体抗铝性强于来自北方石灰性土壤的菌株,这表明外生菌根真菌的铝耐受能力与其原始生境有着密切的联系。外生菌根真菌能分泌多种有机酸,且不同菌株分泌的有机酸种类不同。其中,受铝胁迫分泌量增加最多的是草酸。研究中,铝胁迫能增加大多数铝抗性菌株的草酸分泌量,其中铝抗性最强的Sl 08表现最为明显。但铝胁迫并没有促进具备一定铝抗性的Bo 11和Sl 01草酸的分泌量,同时在铝敏感的菌株中均观察到了草酸分泌量的增加。这表明分泌草酸可能并不是外生菌根真菌抵抗铝毒的唯一途径。对各菌株铝胁迫下对氮,磷及钾的吸收研究表明,除铝敏感菌株Sl 14外,铝胁迫均能促进各供试菌株对氮,磷或钾的吸收。综上,在一定铝浓度下,一些外生菌根真菌可通过增加草酸分泌来抵御铝毒。此外,铝胁迫下外生菌根真菌还可通过调控氮、磷、钾等营养元素的吸收来抵抗铝毒,即通过增加对营养元素的吸收来增强其在铝胁迫下的生存能力,这可能是其抵御铝胁迫的应激反应之一。     

Decolorization of azo, triphenylmethane and anthraquinone dyes by a newly isolated Trametes sp. SQ01 and its laccase

A white-rot fungus, strain SQ01, was isolated from decayed wood in a temperate forest. The strain was identified as a member of genus Trametes, based on the morphological characteristics and a complete sequence analysis of its 18S rRNA gene and ITS region. Strain SQ01 was capable of decolorizing a variety of synthetic dyes, including azo, triphenylmethane, and anthraquinone dyes, with an optimal efficiency of decolorization obtained when dyes added after 5 days of culture, with the exception of Cresol Red, showing that the point of dye addition was an important influencing factor for decolorization by this fungus. All of the tested dyes were decolorized by the purified laccase in the absence of any redox mediators, but only a few were completely removed, while others were not completely degraded even with increased decolorization time.     

Azospirillum genomes reveal transition of bacteria from aquatic to terrestrial environments

Fossil records indicate that life appeared in marine environments ～3.5 billion years ago (Gyr) and transitioned to terrestrial ecosystems nearly 2.5 Gyr. Sequence analysis suggests that "hydrobacteria" and "terrabacteria" might have diverged as early as 3 Gyr. Bacteria of the genus Azospirillum are associated with roots of terrestrial plants; however, virtually all their close relatives are aquatic. We obtained genome sequences of two Azospirillum species and analyzed their gene origins. While most Azospirillum house-keeping genes have orthologs in its close aquatic relatives, this lineage has obtained nearly half of its genome from terrestrial organisms. The majority of genes encoding functions critical for association with plants are among horizontally transferred genes. Our results show that transition of some aquatic bacteria to terrestrial habitats occurred much later than the suggested initial divergence of hydro- and terrabacterial clades. The birth of the genus Azospirillum approximately coincided with the emergence of vascular plants on land.     

Mechanisms and pathways of aniline elimination from aquatic environments

The fate of aniline, a representative of arylamine pollutants derived from the manufacture of dyes, coal liquefaction, and pesticide degradation, was comprehensively evaluated by use of unpolluted and polluted pond water as model environments. Evaporation plus autoxidation proved to be minor elimination mechanisms, removing ca. 1% of the added aniline per day. Instantaneous binding to humic components of a 0.1% sewage sludge inoculum removed 4%. Biodegradation of aniline in pond water was accelerated by the sewage sludge inoculum. A substantial portion of the degraded aniline carbon was mineralized to CO2 within a 1-week period, and microbial biomass was formed as a result of aniline utilization. Biodegradation was clearly the most significant removal mechanism of polluting aniline from pond water. A gas chromatographic-mass spectrometric analysis of biodegradation intermediates revealed that the major pathway of aniline biodegradation in pond water involved oxidative deamination to catechol, which was further metabolized through cis,cis-muconic, beta-ketoadipic, levulinic, and succinic acid intermediates to CO2. Minor biodegradation pathways involved reversible acylation to acetanilide and formanilide, whereas N-oxidation resulted in small amounts of oligomeric condensation products.     

Emission of hydrogen from deep and shallow freshwater environments

In-situ partial pressures of hydrogen in anoxic profundal lake sediments reached values of up to 5 Pa which were more than 5 orders of magnitude lower than the partial pressures of methane. Analysis of gas bubbles collected from anoxic submerged paddy soil showed H₂ partial pressures in the range of 1.8 ± 1.3 Pa being ca. 4 orders of magnitude lower than the CH₄ partial pressures. H₂ emission rates, on the other hand, were less than 3 orders of magnitude lower than the CH₄ emission rates indicating that H₂ and CH₄ were oxidized to a different extent in the rhizosphere of the soil before they reached the atmosphere, or that H₂ was produced by the plants. More than 70% of the emitted H₂ reached the atmosphere via plant-mediated flux. The rest was emitted via ebullition from the anoxic soil and, in addition, was produced in the paddy water. A significant amount of H₂ was indeed found to be produced in the water under conditions where thallic algae and submerged parts of the rice plants produced oxygen by photosynthesis. Very little H₂ was emitted via molecular diffusion through the paddy water; in addition, this amount was less than expected from the degree of supersaturation and the diffusional emission rate of CH₄ indicating a relatively high rate of H₂ consumption in the surface film of the paddy water. The total H₂ source strength of rice paddies and other freshwater environments was estimated to be less than 1 Tg yr^-1, being negligible in the atmospheric budget of H₂.     

Toxicity assessment and microbial degradation of azo dyes

Toxic effluents containing azo dyes are discharged from various industries and they adversely affect water resources, soil fertility, aquatic organisms and ecosystem integrity. They pose toxicity (lethal effect, genotoxicity, mutagenicity and carcinogenicity) to aquatic organisms (fish, algae, bacteria, etc.) as well as animals. They are not readily degradable under natural conditions and are typically not removed from waste water by conventional waste water treatment systems. Benzidine based dyes have long been recognized as a human urinary bladder carcinogen and tumorigenic in a variety of laboratory animals. Several microorganisms have been found to decolourize, transform and even to completely mineralize azo dyes. A mixed culture of two Pseudomonas strains efficiently degraded mixture of 3-chlorobenzoate (3-CBA) and phenol/cresols. Azoreductases of different microorganisms are useful for the development of biodegradation systems as they catalyze reductive cleavage of azo groups (-N=N-) under mild conditions. In this review, toxic impacts of dyeing factory effluents on plants, fishes, and environment, and plausible bioremediation strategies for removal of azo dyes have been discussed.