Schoenoplectus americanus as a potential phytoremediator: in vitro assessment of its ability to remove contaminants in domestic and tannery wastewater.

Growing industrialization and urbanization have led to increased water pollution due to the inadequate treatment and disposal of domestic wastewater (DW) and wastewater produced by industries such as tanneries (TW). These wastewaters are characterized by high concentrations of organic matter, nutrients, sulphates, chlorides and high microbial load. TW also contains phenols and chromium, which disturb and harm the ecosystem the local. The decontamination of wastewater prior to their discharge through biological tools, especially the use of species that are native to the site in need of treatment, has been described as effective and advantageous. This study evaluated the ability of Schoenoplectus americanus, a native plant species from Cordoba (Argentina), to phytoremediate local DW and TW samples at a laboratory scale. The aim was to ascertain whether this system could potentially be considered for the remediation of wastewater in real-world scenarios. S. americanus was able to tolerate pure DW and a 1/20 (v/v) dilution of TW for 30 days under hydroponic conditions. Removal rates ranging from 50% to 89% were obtained for residual organic matter (determined as chemical oxygen demand or COD), total nitrogen (TN) and total phosphorus (TP). Significant removal of total chromium (TCr) and total phenols (TPhs) was also observed in TW (85% and 98%). The number of total coliforms (TC), was reduced by about 96% and 99%. These results indicate that S. americanus is a good candidate for the phytoremediation of regional DW and TW. For this reason, it may be considered for full-scale applications in the future.

Sequential application of activated sludge and phytoremediation with aquatic macrophytes on tannery effluents: in search of a complete treatment.

Tannery effluents with a high organic matter load (indicated by their COD level) have to be treated before they are discharged, so as to minimize their negative impact on the environment. Using field mesocosm systems, this study evaluated the feasibility of treating such effluents through bioaugmentation with activated sludge, followed by phytoremediation with aquatic macrophytes (Lemnoideae subfamily). Regardless of its quality, the activated sludge was able to remove approximately 77% of the COD from effluents with a low initial organic load (up to 1500 mg/L). The macrophytes then enhanced removal (up to 86%), so the final COD values were permissible under the current legislation for effluent discharge. When the initial organic load in the undiluted effluents was higher (around 3000 mg/L), the COD values obtained after consecutive bioaugmentation and phytoremediation were close to the legally allowed limits (583 mg/L), which highlights the potential of phytoremediation as a tertiary treatment. This treatment also brought total coliform counts down to legally acceptable values, without plant biomass decreasing over time. Moreover, the plant biomass remained viable and capable of high COD removal efficiency (around 75%) throughout two additional reuse cycles. These findings indicate that the efficiency of the biological treatments assayed here depends largely on the initial organic load in the tannery effluents. In any case, the sequential application of activated sludge and aquatic macrophytes proved to be a successful alternative for remediation.

Application of two bioassays as potential indicators of phenol phytoremediation efficiency by tobacco hairy roots.

Phenol is one of the contaminants most frequently found in the environment and it is considered a priority pollutant due to their toxic effects. Hairy roots (HR) constitute a good model tool for the removal of this contaminant. In this work, phenol removal using wild type (WT) and double transgenic (DT) Nicotiana tabacum HR was performed with high efficiency (60-80%, for 25-250 mg L-1 phenol solutions, respectively). After phytoremediation process, the toxicity of post removal solutions (PRS) was evaluated through two-toxicity test belonging to two trophic levels, Lactuca sativa test and Rhinella arenarum (AMPHITOX). Toxicity of PRS showed variable results since these solutions were less toxic to L. sativa seeds compared to R. arenarum embryos, which could be attributed to different sensitivities of the exposed organisms. Although PRS obtained using WT and DT HR reduced phenol phytotoxicity on L. sativa seeds, WT PRS were even less toxic than DT PRS according to this test. Regarding AMPHITOX, HR culture medium without phenol but incubated with HR and phenol PRS exerted a toxic effect on the embryos, which could be related to the presence of toxic products derived from HR metabolism. The results demonstrated that an efficient phenol removal is not always accompanied by a considerable reduction of the solution toxicity and therefore, the use of organisms from different trophic levels to evaluate the toxicity after the removal process gains importance.

Deepening the knowledge on the removal of Cr(VI) by L. minuta Kunth: removal efficiency and mechanisms, lipid signaling pathways, antioxidant response, and toxic effects.

Lemna minuta Kunth was used to remove Cr(VI) from aqueous solutions, and some of the mechanisms involved in this process were analyzed. In addition, the cellular signaling mediated by phospholipase D activity as well as antioxidant responses was also evaluated during the process. Cr(VI) removal efficiencies were 40% for 0.5 mg/L, after 24 h, and up to 18% at metal concentrations as high as 5 mg/L. Removal mechanisms displayed by these macrophytes include bioadsorption to cell surfaces and, to a greater extent, Cr internalization and bioaccumulation within cells. Inside of them, Cr(VI) was reduced to Cr(III), a less toxic form of this metal. At the first hours of Cr(VI) exposure, plants were able to sense chromium, activating membrane signal transduction pathways mediated by phospholipase D and phosphatidic acid. Moreover, an increase in the activity of antioxidant enzymes such as superoxide dismutases and peroxidases was observed in the same time. These and other components of the antioxidant defense system would help to reduce the stress generated by the metal. The toxicity of the products formed during the removal process was assessed through Lactuca sativa L. and AMPHIAGU test. It was evidenced that Cr(VI) phytoremediation process by L. minuta plants did not generate acute toxicity neither for L. sativa seeds nor for embryos of Rhinella arenarum (Hensel, 1876). Thus, L. minuta plants could be considered as valuable species for the treatment of waters contaminated with Cr(VI).

Laboratory and field microcosms as useful experimental systems to study the bioaugmentation treatment of tannery effluents.

Tannery effluents require effective treatment prior to their final disposal, and the use of native bacterial consortia could be an appropriate strategy for this purpose. In the present work, consortium SFC 500-1 was found to be highly tolerant to different metals, metalloids and aromatic compounds like phenols. It was also able to remove the black dye commonly used in tanneries. Moreover, it promoted a significant reduction in chemical oxygen demand and exhibited high capability for the simultaneous removal of Cr(VI) and phenol. However, the effectiveness of the remediation processes markedly varied from one experimental system (Erlenmeyer flasks) to another (field microcosm system), highlighting the importance of moving from a small-scale study system to one involving more realistic environmental scenarios. In addition, we found a decrease in the toxicity of the effluent treated with consortium SFC 500-1. Taken together, our results indicate that this consortium possesses great potential for the treatment of tannery effluents. We conclude that for the development of a bioremediation strategy, it is necessary to develop experiments at a larger scale under conditions similar to those of the original system, in order to complete the scenario first created by in vitro approaches.

Synergistic effect of chickpea plants and Mesorhizobium as a natural system for chromium phytoremediation.

The presence of chromium in soils not only affects the physiological processes of plants but also the microbial rhizosphere composition and metabolic activities of microorganisms. Hence, the inoculation of plants with Cr(VI)-tolerant rhizospheric microorganisms as an alternative to reduce Cr phytotoxicity was studied. In this work, chickpea germination was reduced by Cr(VI) concentrations of 150 and 250 mg/L (6 and 33%, respectively); however lower Cr(VI) concentrations negatively affected the biomass. On the other hand, its symbiont, Mesorhizobium ciceri, was able to grow and remove different Cr(VI) concentrations (5-20 mg/L). The inoculation of chickpea plants with this strain exposed to Cr(VI) showed a significantly enhanced plant growth. In addition, inoculated plants accumulated higher Cr concentration in roots than those noninoculated. It is important to note that Cr was not translocated to shoots independently of inoculation. These results suggest that Mesorhizobium's capability to remove Cr(VI) could be exploited for bioremediation. Moreover, chickpea plants would represent a natural system for phytoremediation or phytostabilization of Cr in situ that could be improved with M. ciceri inoculation. This strategy would be considered as a phytoremediation tool with great economic and ecological relevance.

Biotechnological tools to improve bioremediation of phenol by Acinetobacter sp. RTE1.4.

The use of native bacteria is a useful strategy to decontaminate industrial effluents as well as the environment. Acinetobacter sp. RTE1.4 was previously isolated from polluted environments and constitutes a promising alternative for this purpose due to its capability to remove phenol from synthetic solutions and industrial effluents. In this work, this strain was identified at species level as A. tandoii RTE1.4. Phenol degradation pathway was studied and some reaction intermediates were detected, confirming that this strain degraded phenol through ortho-cleavage of the aromatic ring. Phenol removal assays were carried out in a stirred tank bioreactor and a complete degradation of the contaminant was achieved after only 7 h, at an aeration rate of 3 vvm and at agitation of 600 rpm. Moreover, this bacterium was immobilized into calcium alginate beads and an increase in phenol biodegradation with respect to free cells was observed. The immobilized cells were reused for four consecutive cycles and stored at 4°C for 9 months, during which phenol removal efficiency was maintained. Post-removal solutions were evaluated by Microtox® test, showing a toxicity reduction after bacterial treatment. These findings demonstrated that A. tandoii RTE1.4 might be considered as a useful biotechnological tool for an efficient treatment of different solutions contaminated with phenol in bioreactors, using either free or immobilized cells.

Chromium (VI) remediation by a native strain: effect of environmental conditions and removal mechanisms involved.

A native bacterial strain with high capability for Cr (VI) removal was isolated from tannery sediments located in Elena (Córdoba Province, Argentina). The strain was characterized by amplification of 16S rRNA gene and identified as Serratia sp. C8. It was able to efficiently remove different Cr (VI) concentrations in a wide range of pHs and temperatures. The addition of different carbon sources as well as initial inoculum concentration were analyzed, demonstrating that Serratia sp. C8 could reduce 80 % of 20 mg/L Cr (VI) in a medium containing glucose 1 g/L, at pH 6-7 and 28 °C as optimal conditions, using 5 % inoculum concentration. The mechanisms involved in Cr (VI) removal were also evaluated. The strain was capable of biosorpting around 7.5-8.5 % of 20 mg/L Cr on its cell surface and to reduce Cr (VI). In addition, approximately a 54 and 46 % of total Cr was detected in the biomass and in the culture medium, respectively, and in the culture medium, Cr (III) was the predominant species. In conclusion, Serratia sp. C8 removed Cr (VI) and the mechanisms involved in decreasing order of contribution were as follows: reduction catalyzed by intracellular enzymes, accumulation into the cells, and biosorption to the microbial biomass. This strain could be a suitable microorganism for Cr (VI) bioremediation of tannery sediments and effluents or even for other environments contaminated with Cr.

Application of two bacterial strains for wastewater bioremediation and assessment of phenolics biodegradation.

The use of native bacteria is a useful strategy to decontaminate industrial effluents. In this work, two bacterial strains isolated from polluted environments constitutes a promising alternative since they were able to remove several phenolic compounds not only from synthetic solutions but also from effluents derived from a chemical industry and a tannery which are complex matrices. Acinetobacter sp. RTE 1.4 showed ability to completely remove 2-methoxyphenol (1000 mg/L) while Rhodococcus sp. CS 1 not only degrade the same concentration of this compound but also removed 4- chlorophenol, 2,4-dichlorophenol and pentachlorophenol with high efficiency. Moreover, both bacteria degraded phenols naturally present or even exogenously added at high concentrations in effluents from the chemical industry and a tannery in short time (up to 5 d). In addition, a significant reduction of biological oxygen demand and chemical oxygen demand values was achieved after 7 d of treatment for both effluents using Acinetobacter sp. RTE 1.4 and Rhodococcus sp. CS1, respectively. These results showed that Acinetobacter sp. RTE1.4 and Rhodococcus sp. CS 1 might be considered as useful biotechnological tools for an efficient treatment of different effluents, since they showed wide versatility to detoxify these complex matrices, even supplemented with high phenol concentrations.

Characterization of a phenol-degrading bacterium isolated from an industrial effluent and its potential application for bioremediation.

The use of native microorganisms is a useful strategy for phenol bioremediation. In the present work, a bacterial strain, named RTE1.4, was isolated from effluents of a chemical industry. The strain was able to grow at high concentrations of phenol and its derivatives, such as guaiacol, 2,4-dichlorophenol and pentachlorophenol, as well as in a medium containing industrial effluents. This bacterium was identified as Acinetobacter sp. using morphological, physiological, biochemical and 16S rRNA gene analysis. Acinetobacter sp. RTE1.4 degraded phenol (200 to 600 mg/L) at wide pH range and temperature (5-9 and 25-37 degrees C, respectively) demonstrating high adaptation ability to different conditions. The strain would metabolize phenol by the ortho-pathway since catechol 1,2-dioxygenase activity was detected. When bacteria were grown in medium containing phenol, an altered whole-cell protein pattern was observed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), with the lack of some low-molecular mass polypeptides and an increase in the relative abundance of high-molecular mass proteins after treatment. Considering that the use of native strains in bioremediation studies shows several ecological advantages and that the studied bacterium showed high tolerance and biodegradation capabilities, Acinetobacter sp. RTE1.4 could be an appropriate microorganism for improving bioremediation and biotreatment of areas polluted with phenol and/or some of its derivatives. Moreover, the establishment of the optimal growth conditions (pH, temperature, concentration of the pollutant) would provide baseline data for bulk production of the strain and its use in bioremediation processes.

Brassica napus hairy roots and rhizobacteria for phenolic compounds removal.

Phenolic compounds are contaminants frequently found in water and soils. In the last years, some technologies such as phytoremediation have emerged to remediate contaminated sites. Plants alone are unable to completely degrade some pollutants; therefore, their association with rhizospheric bacteria has been proposed to increase phytoremediation potential, an approach called rhizoremediation. In this work, the ability of two rhizobacteria, Burkholderia kururiensis KP 23 and Agrobacterium rhizogenes LBA 9402, to tolerate and degrade phenolic compounds was evaluated. Both microorganisms were capable of tolerating high concentrations of phenol, 2,4-dichlorophenol (2,4-DCP), guaiacol, or pentachlorophenol (PCP), and degrading different concentrations of phenol and 2,4-DCP. Association of these bacterial strains with B. napus hairy roots, as model plant system, showed that the presence of both rhizospheric microorganisms, along with B. napus hairy roots, enhanced phenol degradation compared to B. napus hairy roots alone. These findings are interesting for future applications of these strains in phenol rhizoremediation processes, with whole plants, providing an efficient, economic, and sustainable remediation technology.

Isolation and characterization of a Rhodococcus strain with phenol-degrading ability and its potential use for tannery effluent biotreatment.

INTRODUCTION: Wastewater derived from leather production may contain phenols, which are highly toxic, and their degradation could be possible through bioremediation technologies. MATERIALS, METHODS AND RESULTS: In the present work, microbial degradation of phenol was studied using a tolerant bacterial strain, named CS1, isolated from tannery sediments. This strain was able to survive in the presence of phenol at concentrations of up to 1,000 mg/L. On the basis of morphological and biochemical properties, 16S rRNA gene sequencing, and phylogenetic analysis, the isolated strain was identified as Rhodococcus sp. Phenol removal was evaluated at a lab-scale in Erlenmeyer flasks and at a bioreactor scale in a stirred tank reactor. Rhodococcus sp. CS1 was able to completely remove phenol in a range of 200 to 1,000 mg/L in mineral medium at 30 ± 2 °C and pH 7 as optimal conditions. In the stirred tank bioreactor, we studied the effect of some parameters, such as agitation (200-600 rpm) and aeration (1-3 vvm), on growth and phenol removal efficiency. Faster phenol biodegradation was obtained in the bioreactor than in Erlenmeyer flasks, and maximum phenol removal was achieved at 400 rpm and 1 vvm in only 12 h. Furthermore, Rhodococcus sp. CS1 strain was able to grow and completely degrade phenols from tannery effluents after 9 h of incubation. CONCLUSION: Based on these results, Rhodococcus sp. CS1 could be an appropriate microorganism for bioremediation of tannery effluents or other phenol-containing wastewaters.

Phytoremediation of 2,4-dichlorophenol using wild type and transgenic tobacco plants.

INTRODUCTION: Transgenic plant strategies based on peroxidase expression or overexpression would be useful for phenolic compound removal since these enzymes play an important role in phenolic polymerizing reactions. MATERIAL AND METHODS: Thus, double transgenic (DT) plants for basic peroxidases were obtained and characterized in order to compare the tolerance and efficiency for 2,4-dichlorophenol (2,4-DCP) removal with WT and simple transgenic plants expressing TPX1 or TPX2 gene. Several DT plants showed the expression of both transgenes and proteins, as well as increased peroxidase activity. RESULTS: DT lines showed higher tolerance to 2,4-DCP at early stage of development since their germination index was higher than that of WT seedlings exposed to 25 mg/L of the pollutant. High 2,4-DCP removal efficiencies were found for WT tobacco plants. TPX1 transgenic plants and DT (line d) reached slightly higher removal efficiencies for 10 mg/L of 2,4-DCP than WT plants, while DT plants (line A) showed the highest removal efficiencies (98%). These plants showed an increase of 21% and 14% in 2,4-DCP removal efficiency for solutions containing 10 and 25 mg/L 2,4-DCP, respectively, compared with WT plants. In addition, an almost complete toxicity reduction of postremoval solutions using WT and DT plants was obtained through AMPHITOX test, which indicates that the 2,4-DCP degradation products would be similar for both plants. CONCLUSION: These results are relevant in the field of phytoremediation application and, moreover, they highlight the safety of using DT tobacco plants because nontoxic products were formed after an efficient 2,4-DCP removal.

Evaluation of phenol detoxification by Brassica napus hairy roots, using Allium cepa test.

INTRODUCTION: Meristematic mitotic cells of Allium cepa constitute an adequate material for cytotoxicity and genotoxicity evaluation of environmental pollutants, such as phenol, which is a contaminant frequently found in several industrial effluents. RESULTS AND DISCUSSION: In the present work, Brassica napus hairy roots (HR) were used for phenol removal assays. The toxicity of post-removal solutions (PRS) and phenol solutions was analyzed. These HR removed the contaminant with high efficiency (100-80% for phenol solutions containing 10-250 mg/L, respectively). Phenol solutions treated with B. napus HR showed a significant reduction of general toxicity compared to untreated phenol solutions, since the IC50 values were 318.39 and 229.02 mg/L, respectively. Moreover, PRS presented lower cytotoxicity and genotoxicity than that found in phenol solutions untreated. The mitotic index (MI) observed in meristematic cells treated with PRS (100 and 250 mg/L of phenol) showed an increase of 35% and 42%, whereas the chromosome aberrations showed a significant decrease. According to these results, B. napus HR cultures could be used for the treatment of solutions contaminated with phenol, since we observed not only high removal efficiency, but also an important reduction of the general toxicity, cytotoxicity, and genotoxicity.