Enzymatic detoxification of azo dyes by a multifarious Bacillus sp. strain MR-1/2-bearing plant growth-promoting characteristics.
ABSTRACT: This study was conducted to elucidate the inherent potential of Bacillus sp. MR-1/2, which was isolated from root zone of maize crop grown on a textile wastewater-irrigated soil. The isolated strain was identified through its ribosomal RNA sequence. Under in vitro conditions, the strain demonstrated its tolerance for high concentrations of various heavy metal ions as determined by minimum inhibitory concentration. Moreover, the strain MR-1/2 exhibited many important phytobeneficial traits such as inorganic P solubilization and 1-aminocyclopropane-1-carboxylate (ACC) deaminase ability even under high metal and salt stress. Results showed that the strain proficiently decolorizes various azo dye compounds, e.g., reactive black-5, reactive red-120, and direct blue-1 and congo red, in broth culture. The bioremediation potential of the strain MR-1/2 was further confirmed by analyzing the retrieved azoreductase gene sequence through bioinformatics tools, whereby a subsequent prediction revealed that the azoreductase enzyme activity was involved in decolorization process. When mung bean seeds were grown in pots under various concentrations of decolorized and non-decolorized azo dye, the Bacillus sp. MR-1/2 not only alleviated the azo dye toxicity, but also increased the plant growth parameters. In conclusion, the strain MR-1/2 efficiently decolorized the azo dyes and helped in mung bean plant growth by alleviating azo dye toxicity.
Project description:Textile industry is one of the anthropogenic activities that consume a large amount of water and pollute water bodies. It uses a massive amount of dyes, which is one of the main constituents of polluting textile effluent. In the present research, biodegradation of Acid Blue 113 dye, a commonly used textile di-azo dye, has been studied exploiting <i>Pseudomonas stutzeri</i>, strain AK6. The dye (300 ppm) was decolorized up to 86.2% within 96 h. The metabolites of Acid Blue 113 obtained after biodegradation were identified by various analytical techniques viz. HPLC (high-performance liquid chromatography) and GC-MS (gas chromatography-mass spectrometry). Genome analysis of isolate AK6 using IMG/M (Integrated Microbial Genomes and Microbiomes) system supported the role of azoreductase and laccase for the decolorization and degradation of azo dye. The ability of <i>P. stutzeri</i> AK6 to tolerate high amount of dye makes it a potential candidate for bioremediation and pre-processing to remove dyes from textile effluents.
Project description:The bacterial strain capable of decolorization and detoxification of the Reactive Blue 160 dye was isolated from a dye waste disposal site of Tirupur textile industries. The bacterial strain was screened and selected based on its decolorization capability of RB 160dye, which was identified as Bacillus subtilis by 16S rRNA sequencing. The strain was tested for the decolorization potential under different physio-chemical experimental conditions (pH, temperature, agitation, non-agitation) and observed a complete decolorization at pH 7 and 35?°C under shaking condition within 48?h of time. The enzymes such as, Lignin peroxidase, azoreductase and NADH-DCI were significantly induced in the strain during the decolorization of RB160 dye. Phytotoxicity and microbial toxicity studies revealed that the decolorized product of RB160 dye is less toxic to the plants and microbes. Thus, our results recommend the prospective use of B subtilis in bioremediation of RB160 dye.
Project description:Industrial synthetic dyes cause health and environmental problems. This work describes the isolation of 84 bacterial strains from the midgut of the Lasius niger ant and the evaluation of their potential application in dye bioremediation. Strains were identified and classified as judged by rRNA 16S. The most abundant isolates were found to belong to Actinobacteria (49%) and Firmicutes (47.2%). We analyzed the content in laccase, azoreductase and peroxidase activities and their ability to degrade three known dyes (azo, thiazine and anthraquinone) with different chemical structures. Strain Ln26 (identified as Brevibacterium permense) strongly decolorized the three dyes tested at different conditions. Strain Ln78 (Streptomyces ambofaciens) exhibited a high level of activity in the presence of Toluidine Blue (TB). It was determined that 8.5 was the optimal pH for these two strains, the optimal temperature conditions ranged between 22 and 37?°C, and acidic pHs and temperatures around 50?°C caused enzyme inactivation. Finally, the genome of the most promising candidate (Ln26, approximately 4.2?Mb in size) was sequenced. Genes coding for two DyP-type peroxidases, one laccase and one azoreductase were identified and account for the ability of this strain to effectively oxidize a variety of dyes with different chemical structures.
Project description:<i>Lysinibacillus sphaericus</i> D3 cell-immobilized beads in natural gel sodium alginate decolorized the xylidine orange dye 1-(dimethylphenylazo)-2-naphthol-6-sulfonic acid sodium salt in the laboratory. Optimal conditions were selected for decolorization and the products formed were evaluated for toxicity by disc diffusion assay against common marine bacteria which revealed the non-toxic nature of the dye-degraded products. Decolorization of the brightly colored dye to colorless products was measured on an Ultra Violet-Vis spectrophotometer and its biodegradation products monitored on Thin Layer Chromatographic plate and High Performance Liquid Chromatography (HPLC). Finally, the metabolites formed in the decolorized medium were characterized by mass spectrometry. This analysis confirms the conversion of the parent molecule into lower molecular weight aromatic phenols and sulfonic acids as the final products of biotransformation. Based on the results, the probable degradation products of xylidine orange were naphthol, naphthylamine-6-sulfonic acid, 2-6-dihydroxynaphthalene, and <i>bis</i>-dinaphthylether. Thus, it may be concluded that the degradation pathway of the dye involved (a) reduction of its azo group by azoreductase enzyme (b) dimerization of the hydrazo compound followed by (c) degradation of monohydrazo as well as dimeric metabolites into low molecular weight aromatics. Finally, it may be worth exploring the possibility of commercially utilizing <i>L. sphaericus</i> D3 for industrial applications for treating large-scale dye waste water.
Project description:Release of textile azo dyes to the environment is an issue of health concern while the use of microorganisms has proved to be the best option for remediation. Thus, in the present study, a bacterial consortium consisting of Providencia rettgeri strain HSL1 and Pseudomonas sp. SUK1 has been investigated for degradation and detoxification of structurally different azo dyes. The consortium showed 98-99 % decolorization of all the selected azo dyes viz. Reactive Black 5 (RB 5), Reactive Orange 16 (RO 16), Disperse Red 78 (DR 78) and Direct Red 81 (DR 81) within 12 to 30 h at 100 mg L(-1) concentration at 30 ± 0.2 °C under microaerophilic, sequential aerobic/microaerophilic and microaerophilic/aerobic processes. However, decolorization under microaerophilic conditions viz. RB 5 (0.26 mM), RO 16 (0.18 mM), DR 78 (0.20 mM) and DR 81 (0.23 mM) and sequential aerobic/microaerophilic processes viz. RB 5 (0.08 mM), RO 16 (0.06 mM), DR 78 (0.07 mM) and DR 81 (0.09 mM) resulted into the formation of aromatic amines. In distinction, sequential microaerophilic/ aerobic process doesn't show the formation of amines. Additionally, 62-72 % reduction in total organic carbon content was observed in all the dyes decolorized broths under sequential microaerophilic/aerobic processes suggesting the efficacy of method in mineralization of dyes. Notable induction within the levels of azoreductase and NADH-DCIP reductase (97 and 229 % for RB 5, 55 and 160 % for RO 16, 63 and 196 % for DR 78, 108 and 258 % for DR 81) observed under sequential microaerophilic/aerobic processes suggested their critical involvements in the initial breakdown of azo bonds, whereas, a slight increase in the levels of laccase and veratryl alcohol oxidase confirmed subsequent oxidation of formed amines. Also, the acute toxicity assay with Daphnia magna revealed the nontoxic nature of the dye-degraded metabolites under sequential microaerophilic/aerobic processes. As biodegradation under sequential microaerophilic/aerobic process completely detoxified all the selected textile azo dyes, further efforts should be made to implement such methods for large scale dye wastewater treatment technologies.
Project description:This study was conducted to determine the ability of a bacterial strain FSS2C to ameliorate growth of wheat plants grown under induced stress of reactive black-5 (RB-5). The strain was taxonomically identified as <i>Bacillus firmus</i> on the basis of its 16S rRNA gene sequence analysis. The <i>B. firmus</i> FSS2C was found physiologically potent in phosphate solubilization, indole-3-acetic acid production and ammonia synthesis in the presence of varying concentrations of azo dye RB-5. Moreover, it decolorized RB-5 in vitro with the maximum decolorization (%) found at pH 7 and 30 °C. Inoculation of wheat plants, growing under stress induced by RB-5 dye, with rifampicin-resistant derivatives of the strain FSS2C substantially reduced the cellular oxidative stress, thereby resulting in higher plant biomass as compared to non-inoculated plants. Similarly, the inoculated plants revealed higher nutrient content in shoots as compared to non-inoculated ones. It was concluded that <i>B. firmus</i> strain FSS2C alleviated the oxidative stress impairment caused by reactive black-5 in wheat plants. Therefore, the strain can be used as bio-inoculant in wastewater irrigated soils.
Project description:Azo dyes have become a threat to public health because of its toxicity and carcinogenicity. Azoreductase enzyme plays a pivotal role in the degradation of azodyes released by industrial effluents and other resources. The degradation pathway has to be studied in detail for increasing the activity of azoreductase and for better degradation of azo dyes. But the data available on cyanobacterial azoreductase enzyme and its degradation pathway are still very less. Therefore the present work explored the azoreductase pathway of the cyanobacterium Nostoc sp. PCC7120 for better understanding of the degradation pathway and the other accessory interacting proteins involved. The accessory interacting proteins of azoreductase from cyanobacterium Nostoc sp. PCC7120 were obtained from STRING database. The proteins do not have a comprehensive three dimensional structure and are hypothetical. The secondary structure and functional analysis indicated that the proteins are all soluble proteins, without disulphide bonds and have alpha helices only. The structural prediction and docking study showed that alr2106, alr1063 and alr2326 have best docking result which tally with the STRING database confidence score and thus these proteins could possibly enhance the azoreductase activity and better dye degradation. These results will pave way for further increase in azoreductase activity and for better understanding of the dye degradation pathway.
Project description:Estuaries and coastal environments are often regarded as a critical resource for the bioremediation of organic pollutants such as azo dyes due to their high abundance and diversity of extremophiles. Bioremediation through the activities of azoreductase, laccase, and other associated enzymes plays a critical role in the removal of azo dyes in built and natural environments. However, little is known about the biodegradation genes and azo dye degradation genes residing in sediments from coastal and estuarine environments. In this study, high-throughput sequencing (16S rRNA) of sediment DNA was used to explore the distribution of azo-dye degrading bacteria and their functional genes in estuaries and coastal environments. Unlike laccase genes, azoreductase (azoR), and naphthalene degrading genes were ubiquitous in the coastal and estuarine environments. The relative abundances of most functional genes were higher in the summer compared to winter at locations proximal to the mouths of the Hanjiang River and its distributaries. These results suggested inland river discharges influenced the occurrence and abundance of azo dye degrading genes in the nearshore environments. Furthermore, the azoR genes had a significant negative relationship with total organic carbon, Hg, and Cr (p < 0.05). This study provides critical insights into the biodegradation potential of indigenous microbial communities in nearshore environments and the influence of environmental factors on microbial structure, composition, and function which is essential for the development of technologies for bioremediation in azo dye contaminated sites.
Project description:Biodegradation of Navy N5RL1, a widely used acidic azo dye in carpet industry, was studied by bacterial strain isolated from the dye-contaminated soil collected from a carpet industry premises located in Bhadohi, Sant Ravidas Nagar and Uttar Pradesh, India. The isolated strain was identified as Staphylococcus saprophyticus BHUSS X3 on the basis of morphological, biochemical and 16S rRNA gene sequencing analysis. The strain BHUSS X3 decolorized 95.7 % of dye (100 mg/l) within 6 h at optimum pH 8, temperature 35 °C, inoculum 4.0 % under static condition during 24 h incubation. The isolated bacterial strain BHUSS X3 can toralate dye concentration upto 1,000 mg/l. The dye degradation metabolites were confirmed by analysis of degraded products using UV-Vis spectrophotometric, HPLC and FTIR technique. The phytotoxicity analysis was also conducted on Phaseolus aureus and enhanced seed germination was recorded.
Project description:The gut microbiota enriches the human gene pool and contributes to xenobiotic metabolism. Microbial azoreductases modulate the reduction of azo-bonds, activating produgs and azo polymer-coated dosage forms, or degrading food additives. Here, we aimed to screen the healthy human gut microbiota for food colorant-reducing activity and to characterize factors modulating it. Four representative isolates from screened fecal samples were identified as E. coli (AZO-Ec), E. faecalis (AZO-Ef), E. avium (AZO-Ev) and B. cereus (AZO-Bc). Both AZO-Ef and AZO-Ev decolorized amaranth aerobically and microaerophilically while AZO-Ec and AZO-Bc had higher aerobic reduction rates. The isolates varied in their activities against different dyes, and the azo-reduction activity mostly followed zero-order reaction kinetics, with a few exceptions. Additionally, the isolates had different pH dependence, e.g., AZO-Ec was not affected by pH variation while AZO-Bc exhibited variable degradation kinetics at different pH levels. Cell-free extracts showed NADH-dependent enzymatic activities 14-19 times higher than extracellular fractions. FMN did not affect the reducing activity of AZO-Ef cell-free extract, whereas AZO-Ec, AZO-Ev and AZO-Bc had significantly higher reduction rates in its presence (P?values?=?0.02, 0.0001 and 0.02, respectively). Using Degenerate primers allowed the amplification of azoreductase genes, whose sequences were 98-99% similar to genes encoding FMN-dependent-NADH azoreductases.