The influence of association of plant growth-promoting rhizobacteria and zero-valent iron nanoparticles on removal of antimony from soil by Trifolium repens

Using association of plants, nanomaterials, and plant growth-promoting bacteria (PGPR) is a novel approach in remediation of heavy metal-contaminated soils. Co-application of nanoscale zerovalent iron (nZVI) and PGPR to promote phytoremediation of Sb-contaminated soil was investigated in this study. Seedlings of Trifolium repens were exposed to different regimes of nZVI (0, 150, 300, 500, and 1000 mg/kg) and the PGPR, separately and in combination, to investigate the effects on plant growth, Sb uptake, and accumulation and physiological response of the plant in contaminated soil. Co-application of nZVI and PGPR had positive effects on plant establishment and growth in contaminated soil. Greater accumulation of Sb in the shoots compared to the roots of T. repens was observed in all treatments. Using nZVI significantly increased accumulation capacity of T. repens for Sb with the greatest accumulation capacity of 3896.4 μg per pot gained in the “PGPR+500 mg/kg nZVI” treatment. Adverse impacts of using 1000 mg/kg nZVI were found on plant growth and phytoremediation performance. Significant beneficial effect of integrated use of nZVI and PGPR on plant photosynthesis was detected. Co-application of nZVI and PGPR could reduce the required amounts of nZVI for successful phytoremediation of metalloid polluted soils. Intelligent uses of plants in accompany with nanomaterials and PGPR have great application prospects in removal of antimony from soil.

     

Assessing the impact of nano- and micro-scale zerovalent iron particles on soil microbial activities: particle reactivity interferes with assay conditions and interpretation of genuine microbial effects

The effects of nano-scale and micro-scale zerovalent iron (nZVI and mZVI) particles on general (dehydrogenase and hydrolase) and specific (ammonia oxidation potential, AOP) activities mediated by the microbial community in an uncontaminated soil were examined. nZVI (diameter 12.5 nm; 10 mg g⁻¹ soil) apparently inhibited AOP and nZVI and mZVI apparently stimulated dehydrogenase activity but had minimal influence on hydrolase activity. Sterile experiments revealed that the apparent inhibition of AOP could not be interpreted as such due to the confounding action of the particles, whereas, the nZVI-enhanced dehydrogenase activity could represent the genuine response of a stimulated microbial population or an artifact of ZVI reactivity. Overall, there was no evidence for negative effects of nZVI or mZVI on the processes studied. When examining the impact of redox active particles such as ZVI on microbial oxidation-reduction reactions, potential confounding effects of the test particles on assay conditions should be considered.     

The impact of zero-valent iron nanoparticles upon soil microbial communities is context dependent

Nanosized zero-valent iron (nZVI) is an effective land remediation tool, but there remains little information regarding its impact upon and interactions with the soil microbial community. nZVI stabilised with sodium carboxymethyl cellulose was applied to soils of three contrasting textures and organic matter contents to determine impacts on soil microbial biomass, phenotypic (phospholipid fatty acid (PLFA)), and functional (multiple substrate-induced respiration (MSIR)) profiles. The nZVI significantly reduced microbial biomass by 29 % but only where soil was amended with 5 % straw. Effects of nZVI on MSIR profiles were only evident in the clay soils and were independent of organic matter content. PLFA profiling indicated that the soil microbial community structure in sandy soils were apparently the most, and clay soils the least, vulnerable to nZVI suggesting a protective effect imparted by clays. Evidence of nZVI bactericidal effects on Gram-negative bacteria and a potential reduction of arbuscular mycorrhizal fungi are presented. Data imply that the impact of nZVI on soil microbial communities is dependent on organic matter content and soil mineral type. Thereby, evaluations of nZVI toxicity on soil microbial communities should consider context. The reduction of AM fungi following nZVI application may have implications for land remediation.     

Solid-state fermentation: tool for bioremediation of adsorbed textile dyestuff on distillery industry waste-yeast biomass using isolated Bacillus cereus strain EBT1

Bioremediation of textile dyestuffs under solid-state fermentation (SSF) using industrial wastes as substrate pose an economically feasible, promising, and eco-friendly alternative. The purpose of this study was to adsorb Red M5B dye, a sample of dyes mixture and a real textile effluent on distillery industry waste-yeast biomass (DIW-YB) and its further bioremediation using Bacillus cereus EBT1 under SSF. Textile dyestuffs were allowed to adsorb on DIW-YB. DIW-YB adsorbed dyestuffs were decolorized under SSF by using B. cereus. Enzyme analysis was carried out to ensure decolorization of Red M5B. Metabolites after dye degradation were analyzed using UV–Vis spectroscopy, FTIR, HPLC, and GC-MS. DIW-YB showed adsorption of Red M5B, dyes mixture and a textile wastewater sample up to 87, 70, and 81 %, respectively. DIW-YB adsorbed Red M5B was decolorized up to 98 % by B. cereus in 36 h. Whereas B. cereus could effectively reduce American Dye Manufacture Institute value from DIW-YB adsorbed mixture of textile dyes and textile wastewater up to 70 and 100 %, respectively. Induction of extracellular enzymes such as laccase and azoreductase suggests their involvement in dye degradation. Repeated utilization of DIW-YB showed consistent adsorption and ADMI removal from textile wastewater up to seven cycles. HPLC and FTIR analysis confirms the biodegradation of Red M5B. GC-MS analysis revealed the formation of new metabolites. B. cereus has potential to bioremediate adsorbed textile dyestuffs on DIW-YB. B. cereus along with DIW-YB showed enhanced decolorization performance in tray bioreactor which suggests its potential for large-scale treatment procedures.     

Evaluation of phenanthrene toxicity on earthworm (Eisenia fetida): An ecotoxicoproteomics approach

The goal of this study was to identify promising new biomarkers of phenanthrene by identifying differentially expressed proteins in Eisenia fetida after exposure to phenanthrene. Extracts of earthworm epithelium collected at days 2, 7, 14, and 28 after phenanthrene exposure were analyzed by two dimensional electrophoresis (2-DE) and quantitative image analysis. Comparing the intensity of protein spots, 36 upregulated proteins and 45 downregulated proteins were found. Some of the downregulated and upregulated proteins were verified by MALDI-TOF/TOF-MS and database searching. Downregulated proteins in response to phenanthrene exposure were involved in glycolysis, energy metabolism, chaperones, proteolysis, protein folding and electron transport. In contrast, oxidation reduction, oxygen transport, defense systems response to pollutant, protein biosynthesis and fatty acid biosynthesis were upregulated in phenanthrene-treated E. fetida. In addition, ATP synthase b subunit, lysenin-related protein 2, lombricine kinase, glyceraldehyde 3-phosphate dehydrogenase, actinbinding protein, and extracellular globin-4 seem to be potential biomarkers since these biomarker were able to low levels (2.5 mg kg⁻¹) of phenanthrene. Our study provides a functional profile of the phenanthrene-responsive proteins in earthworms. The variable levels and trends in these spots could play a potential role as novel biomarkers for monitoring the levels of phenanthrene contamination in soil ecosystems.     

Antioxidant defense gene analysis in <Emphasis Type="Italic">Brassica oleracea</Emphasis> and <Emphasis Type="Italic">Trifolium repens</Emphasis> exposed to Cd and/or Pb

This study focused on the expression analysis of antioxidant defense genes in Brassica oleracea and in Trifolium repens. Plants were exposed for 3, 10, and 56 days in microcosms to a field-collected suburban soil spiked by low concentrations of cadmium and/or lead. In both species, metal accumulations and expression levels of genes encoding proteins involved and/or related to antioxidant defense systems (glutathione transferases, peroxidases, catalases, metallothioneins) were quantified in leaves in order to better understand the detoxification processes involved following exposure to metals. It appeared that strongest gene expression variations in T. repens were observed when plants are exposed to Cd (metallothionein and ascorbate peroxidase upregulations) whereas strongest variations in B. oleracea were observed in case of Cd/Pb co-exposures (metallothionein, glutathione transferase, and peroxidase upregulations). Results also suggest that there is a benefit to use complementary species in order to better apprehend the biological effects in ecotoxicology.     

Decolorization of azo dye reactive black B by Bacillus cereus strain HJ-1

Reactive black B (RBB) is a group of azo dyes that are widely used in the textile industry. In this study, a new microbial strain was isolated from azo dye contaminated river sediment which is capable of degrading RBB. The strain was identified as Bacillus cereus strain HJ-1 by 16S rRNA gene sequences analysis. The optimal conditions for RBB decolorization by B. cereus strain HJ-1 are: 25 °C, pH 8, 1 CMC of triton X-100, 0.15 g L^?1 of added yeast extract, 0.125 g L^?1 of added glucose and static culture. Then the toxicity of RBB on the green algae Chlorella vulgaris was determined. The results showed that the median effective concentration (EC₅₀) of RBB for C. vulgaris is 48 mg L^?1 and toxicity will really decrease after decolorization. In the end, B. cereus strain HJ-1 was amended into the origin river sediment and analyzed the whole microbial community structure of river sediment samples by PCR-DGGE technique. The result showed that B. cereus strain HJ-1 could survive in the river sediment after 12 d of incubation. Based on this work, we hope that these findings could provide some useful information for applying the decolorization of RBB in our environment.     

Effects of perfluorooctanoic acid and perfluorooctane sulfonate on acute toxicity,superoxide dismutase,and cellulase activity in the earthworm <Emphasis Type="Italic">Eisenia fetida</Emphasis>

Perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) are the two best-known perfluorinated chemicals and have received much attention due to their ubiquity in the environment. In the present study, we evaluated the effects of PFOS and PFOA on acute toxicity, superoxide dismutase (SOD), and cellulase activities in Eisenia fetida. The results of acute toxicity testing using a filter paper contact test and a natural field soil test showed that PFOA and PFOS exhibited acute toxicity in earthworms, and the toxic effect of PFOS was greater than that of PFOA. The results also showed that avoidance behavior is a more sensitive and easy operation biomarker than acute toxicity and will give us information for early diagnosis of soil pollution, well before the lethal effect becomes apparent. Subchronic exposure to PFOA or PFOS resulted in changes in SOD and cellulase activities in E. fetida, and SOD activity was more sensitive than cellulase activity during early exposure. Based on these findings, we suggest that avoidance behavior and SOD activity in earthworms are suitable biomarkers for evaluating the toxicity of PFOA- and PFOS-contaminated soils. These results indicate that exposure to PFOA and PFOS has a potential impact on soil animals and their environment.     

The fate of dichlorvos in soil

Experiments were conducted to determine the factors responsible for the loss (adsorption, chemical hydrolysis, microbial degradation, etc.) of dichlorvos (2,2-dichlorovingl $\text{0}$,$\text{0}$-dimethyl phosphate) in soil perfusion systems of Houston Black clay. The rate of disappearance from the perfusate (hence the rate of dichlorvos degradation in soil) was related directly to the presence of $\text{Bacillus}$$\text{cereus}$ in the perfusion system, the pH of the system, and the extent of dichlorvos adsorption. Gas liquid chromatographic analyses of the perfusates showed that dichlorvos disappearance was rapid when $\text{B}$. $\text{cereus}$ was added to a previously sterilized soil perfusion system (50% in 3.9 days). Under sterile conditions, 50% of the added dichlorvos was recovered after 10 days. When $\text{B}$. $\text{cereus}$ was added to a mineral salts medium containing dichlorvos as sole ccrbon source, 49% of the initial dichlorvos concentration was degraded in 4 days. The organism was not capable of utilizing dichlorvos as a sole phosphorus source. Chemical hydrolysis of dichlorvos in aqueous, buffered, soil-free systems showed that hydrolysis did not occur in very acid systems (<pH 3.3), but increased with increasing pH values (26% in 4 days at pH 6.9), and was rapid at pH 9.3 (> 99% in 2 days). The extent of dichlorvos adsorption was determined by comparing the initial loss of dichlorvos in a sterile, soil-free extract solution with the initial loss in a sterile soil perfusion system. The rapid initial disappearance of dichlorvos in the presence of sterilized soil was attributed to soil adsorption of the pesticide. After 10 days both systems contained equal concentrations (50%) of dichlorvos. Non-biological mechanisms accounted for 70% of the total degradation of dichlorvos, while bacterial degradation accounted for 30% in the soil perfusion systems.     

The response of Populus spp. to cadmium stress: Chemical,morphological and proteomics study

Poplar (Populus) species are seen as candidates for removing heavy metal contamination from polluted soil. A bottom-up multidisciplinary approach was utilized to compare the performances of clones 58-861 and Poli (Populus nigra) and A4A, a Populus nigra × Populus deltoides hybrid to Cd toxicity. Qualitative and quantitative differences in their tolerance to Cd exposure and the uptake, accumulation and translocation of Cd were noted following the hydroponic exposure of rooted cuttings to 20 μM CdSO₄ for either 48 h or 14 d. Cadmium was less toxic for the hybrid clone A4A as compared to Poli and 58-861. Cd uptake and root to shoot translocation were determined by AAS, and its compartmentation was analyzed using SEM/EDX. A comparative proteomic approach was utilized to identify changes in proteins expression according to dose and time of exposure. Toxicity to Cd mainly influenced proteins related to general defense, stress response and carbohydrate metabolism.     

Dechlorination of chloroorganics,decolorization, and simultaneous bioremediation of Cr6+ from real tannery effluent employing indigenous Bacillus cereus isolate

A native Bacillus cereus isolate has been employed, for the first time, for simultaneous decolorization, dechlorination of chloroorganics, and Cr⁶⁺ remediation from the real tannery effluent. Most of the physicochemical variables in 3:1 diluted effluent were well above the standard prescribed limits, which decreased substantially upon microbial treatment. The extent of bioremediation was better in diluted (3:1) as compared to undiluted effluent supplemented with nutrients and augmented with B. cereus isolate. Maximum growth, effluent decolorization (42.5 %), dechlorination (74.1 %), and Cr⁶⁺ remediation (34.2 %) were attained with 4.0 % (v/v) inoculum, 0.8 % glucose, and 0.2 % ammonium chloride in 3:1 diluted effluent at natural pH (8.1) within 72 h of incubation. The efficiency of bioremediation in a bioreactor was higher as compared to a flask trial during 72 h of incubation: decolorization (47.9 %) was enhanced by 5.4 %, dechlorination (77.4 %) by 3.3 %, and Cr⁶⁺ removal (41.7 %) by 7.5 % at an initial color of 286 Pt-Co units and initial concentration of 62 mg chloride ions and 108 mg l^?1 Cr⁶⁺. Immobilized biomass of Pseudomonas putida and B. cereus coculture enhanced the extent of Cr⁶⁺ remediation (51.9 %) by 10.2 % compared to the bioreactor trial. Chromate reductase activity and reduced Cr directly correlated and were mainly associated with soluble fraction of B. cereus plus effluent natural microflora. The GC-MS analyses revealed the formation of metabolites such as acetic acid and 2-butenoic acid in bacterially treated effluent. The supplementation of nutrients along with B. cereus augmentation is required for efficient effluent bioremediation.     

Zero-valent iron particles for PCB degradation and an evaluation of their effects on bacteria,plants, and soil organisms

Two types of nano-scale zero-valent iron (nZVI-B prepared by borohydride reduction and nZVI-T produced by thermal reduction of iron oxide nanoparticles in H₂) and a micro-scale ZVI (mZVI) were compared for PCB degradation efficiency in water and soil. In addition, the ecotoxicity of nZVI-B and nZVI-T particles in treated water and soil was evaluated on bacteria, plants, earthworms, and ostracods. All types of nZVI and mZVI were highly efficient in degradation of PCBs in water, but had little degradation effect on PCBs in soil. Although nZVI-B had a significant negative impact on the organisms tested, treatment with nZVI-T showed no negative effect, probably due to surface passivation through controlled oxidation of the nanoparticles.

     

Exploring the potential of applying proteomics for tracking bisphenol A and nonylphenol degradation in activated sludge

A significant percentage of bisphenol A and nonylphenol removal in municipal wastewater treatment plants relies on biodegradation. Nonetheless, incomplete information is available concerning their degradation pathways performed by microbial communities in activated sludge systems. Hydroquinone dioxygenase (HQDO) is a specific degradation marker enzyme, involved in bisphenol A and nonylphenol biodegradation, and it can be produced by axenic cultures of the bacterium Sphingomonas sp. strain TTNP3. Proteomics, a technique based on the analysis of microbial community proteins, was applied to this strain. The bacterium proteome map was obtained and a HQDO subunit was successfully identified. Additionally, the reliability of the applied proteomics protocol was evaluated in activated sludge samples. Proteins belonging to Sphingomonas were searched at decreasing biomass ratios, i.e. serially diluting the bacterium in activated sludge. The protein patterns were compared and Sphingomonas proteins were discriminated against the ones from sludge itself on 2D-gels. The detection limit of the applied protocol was defined as 10^?3 g TTNP3 g^?1 total suspended solids (TSSs). The results proved that proteomics can be a promising methodology to assess the presence of specific enzymes in activated sludge samples, however improvements of its sensitivity are still needed.     

Antibacterial activity of different natural honeys from Transylvania,Romania

Honey is used in food industry and medicine due to its nutritive, therapeutic and dietetic qualities. The microbiological characteristics of 10 unpasteurized honey samples of known origin, collected from Transylvania beekeepers (Romania) were determined. The antibacterial activity of these types of honey against Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Salmonella enteritidis, Salmonella anatum, Salmonella choleraesuis, Bacillus cereus, Bacillus subtilis subsp. spizizenii and Listeria monocytogenes strains was studied. The most sensitive to the antibacterial activity were the two staphylococus strains (the largest diameter of inhibition zone was 18 mm) and B. subtilis strains (13.5 mm). The strains of B. cereus, E. coli, L. monocytogenes and Salmonella spp. were found to present resistance to some of the honey samples. Manna, sunflower and polyfloral honeys presented high antibacterial activity while acacia and linden honeys had a lower activity in terms of the number of sensible strains. Statistical analysis shows that the type of strains and the type of honey have influence on the diameter of inhibition.     

Linking NE1545 gene expression with cell volume changes in Nitrosomonas europaea cells exposed to aromatic hydrocarbons

Nitrosomonas europaea, a model ammonia oxidizing bacterium, was exposed to a wide variety of aromatic hydrocarbons in 3 h batch assays. The expression of NE1545, a phenol sentinel gene involved in fatty acid metabolism, was monitored via quantitative real-time polymerase chain reaction (qRT-PCR) and a Coulter Counter technique was used to monitor changes in cell volume. Decreases in cell volume and NE1545 gene expression correlated strongly with exposure to aromatic hydrocarbons that possessed a single polar group substitution (e.g. phenol and aniline). Aromatic hydrocarbons that contain no polar group substitutions (e.g. toluene) or multiple polar group substitutions (e.g. p-hydroquinone) caused negligible changes in NE1545 expression and cell volume. The oxidation of aromatic hydrocarbons by N. europaea from configurations without a single polar group to one with two polar groups (e.g. p-cresol oxidized to 4-hydroxybenzyl alcohol) and from configurations with no polar groups to one with a single polar group (e.g. ethylbenzene oxidized to 4-ethylphenol) greatly influenced NE1545 gene expression and observed changes in cell volume. Nitrification inhibition in N. europaea by the aromatic hydrocarbons was found to be completely reversible; however, the decreases in cell volume were not reversible suggesting a physical change in cell membrane composition. Ammonia monooxygenase blocking studies showed that the chemical exposure that was responsible for the cell volume decrease and up-regulation in gene expression and not the observed inhibition. N. europaea is the first bacterium shown to experience significant changes in cell volume when exposed to μM concentrations of aromatic hydrocarbons, three orders of magnitude lower than previous studies with other bacteria.     

Simultaneous adsorption and degradation of Zn2+ and Cu2+ from wastewaters using nanoscale zero-valent iron impregnated with clays

Clays such as kaolin, bentonite and zeolite were evaluated as support material for nanoscale zero-valent iron (nZVI) to simultaneously remove Cu²⁺ and Zn²⁺ from aqueous solution. Of the three supported nZVIs, bentonite-supported nZVI (B-nZVI) was most effective in the simultaneous removal of Cu²⁺ and Zn²⁺ from a aqueous solution containing a 100 mg/l of Cu²⁺ and Zn²⁺, where 92.9 % Cu²⁺ and 58.3 % Zn²⁺ were removed. Scanning electronic microscope (SEM) revealed that the aggregation of nZVI decreased as the proportion of bentonite increased due to the good dispersion of nZVI, while energy dispersive spectroscopy (EDS) demonstrated the deposition of copper and zinc on B-nZVI after B-nZVI reacted with Cu²⁺ and Zn²⁺. A kinetics study indicated that removing Cu²⁺ and Zn²⁺ with B-nZVI accorded with the pseudo first-order model. These suggest that simultaneous adsorption of Cu²⁺and Zn²⁺ on bentonite and the degradation of Cu²⁺and Zn²⁺ by nZVI on the bentonite. However, Cu²⁺ removal by B-nZVI was reduced rather than adsorption, while Zn²⁺ removal was main adsorption. Finally, Cu²⁺, Zn²⁺, Ni²⁺, Pb²⁺ and total Cr from various wastewaters were removed by B-nZVI, and reusability of B-nZVI with different treatment was tested, which demonstrates that B-nZVI is a potential material for the removal of heavy metals from wastewaters.     

Heterogeneous Fenton oxidation of Direct Black G in dye effluent using functional kaolin-supported nanoscale zero iron

This study investigated kaolin-supported nanoscale zero-valent iron (nZVI/K) as a heterogeneous Fenton-like catalyst for the adsorption and oxidation of an azo dye, Direct Black G (DBG). New findings suggest that kaolin as a support material not only reduced the aggregation of nanoscale zero-valent iron (nZVI) but also improved the adsorption of DBG. It consequently improved Fenton oxidation by increasing the local concentration of DBG in the vicinity of nZVI. This was confirmed by scanning electron microscopy and X-ray diffraction for the surface morphology of nZVI/K before and after the Fenton-like reaction. Furthermore, nZVI/K proved to be a catalyst for the heterogeneous Fenton-like oxidation of the DBG process in the neutral pH range. More than 87.22 % of DBG was degraded, and 54.60 % of total organic carbon was removed in the optimal conditions: 0.6 g/L dosage of nZVI/K, 33 mM H₂O₂, 100 mg/L initial DBG concentration, temperature of 303 K and pH of 7.06. Finally, it was demonstrated that nZVI/K removed DBG from dye wastewater through the processes of adsorption and oxidation.     

Differential expression profile of membrane proteins in Aplysia pleural–pedal ganglia under the stress of methyl parathion

This study was aimed to analyze the alteration of membrane protein profiles in Aplysia juliana Quoy & Gaimard (A. juliana) pleural–pedal ganglia under MP exposure. Both the results of GC–MS analysis and the activity assay of acetylcholinesterase (AChE), superoxide dismutase (SOD), catalase (CAT) reveal that MP toxicological effects on Aplysia left and right pleural–pedal ganglia are different under 7 and 14 days of exposure. Therefore, Aplysia were subjected for exposure at two concentrations (1 and 2 mg/l) of MP for 7 and 14 days for membrane proteomic study. As a result, 19 and 14 protein spots were differentially expressed in A. juliana left pleural–pedal ganglia under 7 and 14 days treatment, and 20 and 14 protein spots found with differential expressions in their right ganglia under the same treatment, respectively. Several proteins with expression variations were detected from both the left and right pleural–pedal ganglia; however, most proteins have distinctive expressions, indicating different mechanisms might be involved in initiating MP toxicology in left and right ganglia. Among the total differential protein spots obtained, 29 proteins were classed as membrane proteins. These proteins are mainly involved in the metabolism process, cell redox homeostasis, signal transduction, immunology, intracellular transport and catalysis, indicating MP toxicity in mollusks seems to be complex and diverse. Some differentially expressed proteins were further confirmed by Western blotting and quantitative real-time PCR. These results might provide renovated insights to reveal the mechanism of MP-induced neurotoxicity, and the novel candidate biomarkers might have potential application for environmental evaluation of MP pollution level.