Low temperature reduction of hexavalent chromium by a microbial enrichment consortium and a novel strain of Arthrobacter aurescens.
ABSTRACT: Chromium is a transition metal most commonly found in the environment in its trivalent [Cr(III)] and hexavalent [Cr(VI)] forms. The EPA maximum total chromium contaminant level for drinking water is 0.1 mg/l (0.1 ppm). Many water sources, especially underground sources, are at low temperatures (less than or equal to 15 Centigrade) year round. It is important to evaluate the possibility of microbial remediation of Cr(VI) contamination using microorganisms adapted to these low temperatures (psychrophiles).Core samples obtained from a Cr(VI) contaminated aquifer at the Hanford facility in Washington were enriched in Vogel Bonner medium at 10 Centigrade with 0, 25, 50, 100, 200, 400 and 1000 mg/l Cr(VI). The extent of Cr(VI) reduction was evaluated using the diphenyl carbazide assay. Resistance to Cr(VI) up to and including 1000 mg/l Cr(VI) was observed in the consortium experiments. Reduction was slow or not observed at and above 100 mg/l Cr(VI) using the enrichment consortium. Average time to complete reduction of Cr(VI) in the 30 and 60 mg/l Cr(VI) cultures of the consortium was 8 and 17 days, respectively at 10 Centigrade. Lyophilized consortium cells did not demonstrate adsorption of Cr(VI) over a 24 hour period. Successful isolation of a Cr(VI) reducing organism (designated P4) from the consortium was confirmed by 16S rDNA amplification and sequencing. Average time to complete reduction of Cr(VI) at 10 Centigrade in the 25 and 50 mg/l Cr(VI) cultures of the isolate P4 was 3 and 5 days, respectively. The 16S rDNA sequence from isolate P4 identified this organism as a strain of Arthrobacter aurescens, a species that has not previously been shown to be capable of low temperature Cr(VI) reduction.A. aurescens, indigenous to the subsurface, has the potential to be a predominant metal reducer in enhanced, in situ subsurface bioremediation efforts involving Cr(VI) and possibly other heavy metals and radionuclides.
Project description:Hexavalent chromium-resistant Ochrobactrum intermedium BCR400 was isolated from chromium contaminated soil collected from Vadodara, Gujarat. It reduced 100 mg Cr(VI)/L completely in 52 h with initial Cr(VI) reduction rate of 1.98 mg/L/h. The Cr(VI) reduction rate decreased with increase in Cr(VI) concentration from 100 to 500 mg/L. The addition of anthraquinone-2-sulphonic acid (AQS) to culture O. intermedium BCR400 significantly enhanced its chromium reduction rate. The activation energy of AQS-mediated Cr(VI) reduction (120.69 KJ/mol) was 1.1-fold lower than non-mediated Cr(VI) reduction. An increase in the activities of quinone reductase and chromate reductase in cells grown in presence of AQS/AQS + Cr(VI) suggests their role in reduction of Cr(VI) by O. intermedium. Both chromate reductase and quinone reductase activities were FAD independent, required NADH as reductant, displayed maximum activity at pH (7.0) and temperature (30 °C). Thus Cr(VI) bioremediation potential of O. intermedium can be enhanced by augmentation of system with AQS as redox mediator.
Project description:Hexavalent chromium reduction and accumulation by Acinetobacter AB1 isolated from Fez tanneries effluents were tested. The effects of some environmental factors such as pH, temperature, and exposure time on Cr(VI) reduction and resistance were investigated. We found that this strain was able to resist to concentrations as high as 400 mg/l of Cr(VI). Moreover, pH 10 and the temperature 30°C constitute favourable conditions to the growth and reduction of Acinetobacter AB1. Complete reduction of Cr(VI) was observed at low initial Cr(VI) concentrations of 50 mg/l after 72 h of incubation. Furthermore, Transmission electron microscope (TEM) analysis showed morphological changes in AB1 strain due 48H exposure to 100 mg/l chromate concentration and revealed circular electron dense (dark black point) inclusion within the cell cytoplasm suggesting chromium deposition within the cells.
Project description:The current study aimed to isolate and characterize a chromate-resistant bacterium from tannery effluent, able to reduce Cr(VI) aerobically at high pH and salinity. Environmental contamination by hexavalent chromium, Cr(VI), presents a serious public health problem. Enrichment led to the isolation of 12 bacteria displaying different degrees of chromate reduction. Phenotypic characterization and phylogenetic analysis based on 16S rDNA sequence comparison indicated that the most potent strain belonged to the genus Halomonas. The new strain designated as Halomonas sp. M-Cr was able to reduce 82% of 50 mg L(-1) Cr(VI) in 48 h, concomitant with discolouring of yellow colour of the medium and formation of white insoluble precipitate of Cr(III). It exhibited growth up to 3500 mg L(-1) Cr(VI), 20% NaCl and showed strong Cr(VI) reduction under alkaline condition, pH 10. Scanning electron microscopy revealed precipitation of chromium hydroxide on bacterial cell surfaces, which showed characteristic peak of chromium in energy-dispersive X-ray analysis. Plackett-Burman design was used to evaluate the influence of related parameters for enhancing Cr(VI) reduction. Glucose, yeast extract and KH2PO 4 were confirmed as significant variables in the medium. Data suggest Halomonas sp. M-Cr as a promising candidate for bioremediation of Cr(VI) contaminated effluents particularly in saline and alkaline environments. Up to our knowledge, this is the first report on isolation of haloalkaliphilic Halomonas sp. from tannery effluent.
Project description:The bacterium MNU16 was isolated from contaminated soils of coal mine and subsequently screened for different plant growth promoting (PGP) activities. The isolate was further identified by 16S rRNA sequencing as Bacillus subtilis MNU16 with IAA concentration (56.95 ± 0.43 6?g/ml), siderophore unit (9.73 ± 2.05%), phosphate solubilization (285.13 ± 1.05 ?g/ml) and ACC deaminase activity (116.79 ± 0.019 ?moles ?-ketobutyrate/mg/24 h). Further, to evaluate the metal resistance profile of bacterium, the isolate was screened for multi-metal resistance (viz. 900 mg/L for Cr, 600 mg/L for As, 700 mg/L for Ni and 300 mg/L for Hg). Additionally, the resistance pattern of B. subtilis MNU16 against Cr(VI) (from 50 to 300 mg/L) treatments were evaluated. An enriched population was observed at 0-200 mg/L Cr(VI) concentration while slight reductions were observed at 250 and 300 mg/L Cr(VI). Further, the chromium reduction ability at 50 mg/L of Cr(VI) highlighted that the bacterium B. subtilis MNU16 reduced 75% of Cr(VI) to 13.23 mg/L within 72 h. The localization of electron dense precipitates was observed in the TEM images of B. subtilis MNU16 which is might be due to the reduction of Cr(VI) to Cr(III). The data of fluorescence microscopy and flow cytometry with respect to Cr(VI) treatments (50-300 mg/L) showed a similar pattern and clearly revealed the less toxic effect of hexavalent chromium upto 200 mg/L Cr(VI) concentration. However, toxicity effects were more pronounced at 300 mg/L Cr(VI). Therefore, the present study suggests that the plant growth promoting potential and resistance efficacy of B. subtilis MNU16 will go a long way in developing an effective bioremediation approach for Cr(VI) contaminated soils.
Project description:Microbial mats are rarely reported for chromium-polluted ecosystems, hence information on their bacterial diversity and role in chromium removal are very scarce. We investigated the role of nine microbial mats, collected from three quarry sumps of chromium mining sites, in the removal of hexavalent chromium [Cr(VI)]. Bacterial diversity in these mats and community shifts after incubation with Cr(VI) have been investigated using MiSeq sequencing. In nature, a chromium content of 1,911 ± 100 mg kg-1 was measured in the microbial mats, constituting the third highest source of environmentally available chromium. The mats were able to remove 1 mg l-1 of Cr(VI) in 7 days under aerobic conditions. MiSeq sequencing of the original mats yielded 46-99% of the sequences affiliated to Proteobacteria, Firmicutes and Actinobacteria. When the mats were incubated with Cr(VI), the bacterial community shifted in the favor of Alphaproteobacteria and Verrucomicrobiae. We conclude that microbial mats in the quarry sumps harbor diverse microorganisms with the ability to remove toxic Cr(VI), hence these mats can be potentially used to remove chromium from polluted waters.
Project description:Two chromium-resistant bacteria (IFR-2 and IFR-3) capable of reducing/transforming Cr(VI) to Cr(III) were isolated from tannery effluents. Isolates IFR-2 and IFR-3 were identified as Staphylococcus aureus and Pediococcus pentosaceus respectively by 16S rRNA gene sequence analyses. Both isolates can grow well on 2,000 mg/l Cr(VI) (as K(2)Cr(2)O(7)) in Luria-Bertani (LB) medium. Reduction of Cr(VI) was found to be growth-associated in both isolates and IFR-2 and IFR-3 reduced 20 mg/l Cr(VI) completely in 6 and 24 h respectively. The Cr(VI) reduction due to chromate reductase activity was detected in the culture supernatant and cell lysate but not at all in the cell extract supernatant of both isolates. Whole cells of IFR-2 and IFR-3 converted 24 and 30% of the initial Cr(VI) concentration (1 mg/l) in 45 min respectively at 37°C. NiCl(2) stimulated the growth of IFR-2 whereas HgCl(2) and CdCl(2) significantly inhibited the growth of both isolates. Optimum temperature and pH for growth of and Cr(VI) reduction by both isolates were found to be between 35 and 40°C and pH 7.0 to 8.0. The two bacterial isolates can be good candidates for detoxification of Cr(VI) in industrial effluents.
Project description:Identifying and quantifying chromium in water are important for the protection of precious water resources from chromium pollution. Standard methods however are unable to easily distinguish toxic hexavalent chromium, Cr(VI), from innocuous trivalent chromium, Cr(III), are time-consuming, or require large sample quantity. We show in this report that Cr(VI) and Cr(III) in water can be differentiated based on their distinct spectral features of surface-enhanced Raman scattering (SERS). Their SERS signals exhibit different pH dependences: the SERS features of Cr(VI) and Cr(III) are most prominent at pH values of 10 and 5.5, respectively. The obtained limit of detection of Cr(VI) in water is below 0.1 mg/L. Both concentration curves of their SERS signals show Langmuir sorption isotherm behavior. A procedure was developed to quantify Cr(VI) concentration based on the direct retrieval or addition method with an error of 10%. Finally, the SERS detection of Cr(VI) is shown to be insensitive to co-present Cr(III). The developed SERS procedure offers potential to monitor toxic chromium in fields.
Project description:An efficient keratinolytic strain of Stenorophomonas maltophilia KB13 was isolated from feather disposal site of Bilaspur, Chhattisgarh, India. The strain could metabolize 10 g/l chicken feathers as sole source of carbon and nitrogen. Soluble protein, amino acid, and cysteine content were found to be maximum (690.6 ± 8.7, 688.9 ± 9.12 and 21 ± 0.36 µg/ml, respectively) at late logarithmic phase of growth. Protease and keratinase activity reached its maximum level (103.26 ± 7.09 and 178.5 ± 9.10 U/ml) at the 4th day of incubation. The feather protein hydrolysate (FPH) obtained after degradation of chicken feathers was utilized to reduce hexavalent chromium. About 78.4 ± 2.4 and 63.6 ± 2.2 % reduction of 50 and 100 mg/l Cr(VI), respectively, was observed after 60 min of incubation with FPH. Further, there was no effect of autoclaved FPH on Cr(VI) reduction indicating that any bacterial enzyme was not involved in reduction process. Cr(VI) reduction was significantly inhibited by 10 mm Hg2+ ions indicating the role of sulfur-containing amino acids in reduction process. FTIR analysis confirmed that chromium reduction occurred due to oxidation of amino acids cysteine and cystine. This study shows that FPH arising after feather degradation can be employed as a potential candidate for the reduction of hexavalant chromium.
Project description:Contaminated water with hexavalent chromium Cr(VI) is a serious environmental problem. This study aimed to evaluate the Cr(VI) removal by zero valent iron nanoparticles (nZVI) reduction process and the impact of Cr(VI), nZVI and combined treatment with nZVI and Cr(VI) on tomato growth performance. To evaluate the Cr(VI) toxic effect on germination capability, seeds were exposed to increasing Cr(VI) concentrations up to 1000?mg?L-1. The inhibition of seed germination and the decrease of hypocotyl and root length started from Cr(VI) 5?mg?L-1. Under treatment with Cr(VI)?+?nZVI 5?mg?L-1, seed germination, hypocotyl and root length resulted significantly higher compared to Cr(VI) 5?mg?L-1 treatment. The impact of only nZVI was investigated on chlorophyll and carotenoid in leaves; iron levels in leaves, roots, fruits and soil; carotenoid, fat-soluble vitamin and nicotianamine in mature fruits. A significant increase of leaf chlorophyll and carotenoids was observed after nZVI 5?mg?L-1 treatment compared to controls. No significant variations were observed in carotenoids, fat-soluble vitamins and nicotianamine levels after treatment with nZVI 5?mg?L-1 in mature fruits. For their ability to reduce Cr(VI) and to stimulate tomato growth, nZVI might to be considered as alternative for remediation purposes.
Project description:Nanometer zero-valent iron (nZVI) has been widely used in the treatment of heavy metals such as hexavalent chromium (Cr(VI)). A novel composite of bead string-structured nZVI on modified activated carbon (nZVI-MAC) is prepared here, using polyethylene glycol as the stable dispersant rather than traditional ethanol during the loading process. The microstructure characterization shows that nZVI particles are loaded on MAC with a bead string structure in large quantity and stably due to the addition of hydroxyl functional groups on the surface by polyethylene glycol. Experiments on the treatment of Cr(VI) in wastewater show that the reaction process requires only 20 min to achieve equilibrium. The removal rate of Cr(VI) with a low concentration (80-100 mg/L) is over 99% and the maximum saturation removal capacity is up to 66 mg/g. The system converts Cr(VI) to trivalent chromium (Cr(III)) through an oxidation-reduction effect and forms an insoluble material with iron ions by coprecipitation, which is then adsorbed on the surface of the nZVI-MAC. The process conforms to the quasi-second order adsorption kinetics equation (mainly chemical adsorption process).