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Functional effects of the bacterial insecticide Bacillus thuringiensis var. kurstaki on aquatic microbial communities.

Epilithic microbial communities were colonized on leaf disks and exposed to commercial preparations of Bacillus thuringiensis var. kurstaki (Btk) in aquatic microcosms. Responses in terms of microbial respiration, bacterial cell density, protozoan density, and microbial decomposition activity were measured. Test concentrations for treatments with Dipel 64AF and Dipel 8AF in microcosms were the expected environmental concentration (EEC) of 20 IU/ml, 100x the EEC, and 1000x the EEC. Bacterial cell density in the biofilm of leaf disks was significantly increased at concentrations as low as the EEC. There were no concomitant alterations in protozoan density. Microbial respiration was significantly increased, and decomposition activity was significantly decreased, but only at the artificially high concentration of 1000x the EEC. This effect was attributed to the spore-crystal component rather than formulation ingredients. Microbial decomposition of leaf material was also determined in outdoor stream channels treated at concentrations ranging from the EEC to 100x the EEC. Although there tended to be reduced decomposition activity in treated channels, there were no significant differences in mass loss of leaf material between treated and control channels. Various regression, classification, and ordination procedures were applied to the experimental data, and none indicated significant treatment effects. These results from laboratory and controlled field experiments indicate that contamination of watercourses with Btk is unlikely to result in significant adverse effects on microbial community function in terms of detrital decomposition.

Agammaglobulinaemia Tyrosine Kinase↗

Phytoremediation of methylmercury pollution: merB expression in Arabidopsis thaliana confers resistance to organomercurials.

Methylmercury is an environmental toxicant that biomagnifies and causes severe neurological degeneration in animals. It is produced by bacteria in soils and sediments that have been contaminated with mercury. To explore the potential of plants to extract and detoxify this chemical, we engineered a model plant, Arabidopsis thaliana, to express a modified bacterial gene, merBpe, encoding organomercurial lyase (MerB) under control of a plant promoter. MerB catalyzes the protonolysis of the carbon---mercury bond, removing the organic ligand and releasing Hg(II), a less mobile mercury species. Transgenic plants expressing merBpe grew vigorously on a wide range of concentrations of monomethylmercuric chloride and phenylmercuric acetate. Plants lacking the merBpe gene were severely inhibited or died at the same organomercurial concentrations. Six independently isolated transgenic lines produced merBpe mRNA and MerB protein at levels that varied over a 10- to 15-fold range, and even the lowest levels of merBpe expression conferred resistance to organomercurials. Our work suggests that native macrophytes (e.g., trees, shrubs, grasses) engineered to express merBpe may be used to degrade methylmercury at polluted sites and sequester Hg(II) for later removal.

Air Pollutants↗

Environmental fate of the antifouling compound zinc pyrithione in seawater.

To perform a thorough risk assessment of the new antifouling compound zinc pyrithione (ZnPT2), additional information regarding the fate of the compound is needed. The present study examined the recovery and transchelation of ZnPT2 in controlled laboratory experiments, photodegradation experiments, and a fate study in a large-scale field experiment. Chemical analyses were performed using a newly developed method for simultaneous analyses of ZnPT2 and copper pyrithione (CuPT2). Furthermore, two antifouling paints containing ZnPT2 were examined for the fate of leaching biocide. Naturally occurring ligands and metals in seawater influence the stability of ZnPT2. The presence of free Cu2+, which is present naturally in the seawater or is released from copper-containing paints, results in a partial transchelation of ZnPT2 into CuPT2. A complete transchelation of ZnPT2 into CuPT2 was observed when Cu2+ was present at an equimolar concentration in the absence of interfering ligands. When ZnPT2 was leached from antifouling paints containing both ZnPT2 and Cu2O, CuPT2 was found, with no trace of ZnPT2. Photodegradation was low in natural waters and absent from 1 m or more below the surface. The results show that ZnPT2 has a low persistence in seawater when leached from antifouling paints. However, the more stable and toxic transchelation product CuPT2 is formed, which has the potential to accumulate in the sediments and, therefore, should be included in both chemical analysis and risk assessment of ZPT2.

Biodegradation, Environmental↗

Bacterial dehalogenation.

Halogenated organic compounds are produced industrially in large quantities and represent an important class of environmental pollutants. However, an abundance of haloorganic compounds is also produced naturally. Bacteria have evolved several strategies for the enzyme-catalyzed dehalogenation and degradation of both haloaliphatic and haloaromatic compounds: (i) Oxidative dehalogenation is the result of mono- or dioxygenase-catalyzed, co-metabolic or metabolic reactions. (ii) In dehydrohalogenase-catalyzed dehalogenation, halide elimination leads to the formation of a double bond. (iii) Substitutive dehalogenation in most cases is a hydrolytic process, catalyzed by halidohydrolases, but there also is a "thiolytic" mechanism with glutathione as cosubstrate. Dehalogenation by halohydrin hydrogen-halide lyases is the result of an intramolecular substitution reaction. (iv) A distinct dechlorination mechanism involves methyl transfer from chloromethane onto tetrahydrofolate. (v) Reductive dehalogenations are co-metabolic processes, or they are specific reactions involved in substrate utilization (carbon metabolism), or reductive dehalogenation is coupled to energy conservation: some anaerobic bacteria use a specific haloorganic compound as electron acceptor of a respiratory process. This review discusses the mechanisms of enzyme-catalyzed dehalogenation reactions, describes some pathways of the bacterial degradation of haloorganic compounds, and indicates some trends in the biological treatment of organohalogen-polluted air, groundwater, soil, and sediments.

Bacteria↗

Evaluation of bioventing on a gasoline-ethanol contaminated undisturbed residual soil.

Remediation methods for environmental contamination problems based on physical or chemical processes frequently present low efficiency and/or high costs. On the other hand, biological treatment is being proved to be an accessible alternative for soil and water remediation. Bioventing is commonly used for petroleum hydrocarbon (PHC) spills. This process provides better subsurface oxygenation, thus stimulating degradation by indigenous microorganisms. In Brazil, gasoline and ethanol are routinely mixed; some authors suggest that despite gasoline high degradability, its degradation in the aquifer is hindered by the presence of much rapidly degrading ethanol. The present study evaluates a bioventing treatment of a gasoline-ethanol contaminated undisturbed residual soil from Rio de Janeiro. Contamination and treatment effects were monitored by conventional microbiology methods, chemical analysis, and ground penetrating radar (GPR) measurements. Results of culturable bacterial population counts show the effect of contamination and bioventing on the microbiota of gasoline and gasoline-ethanol containing soils; however, GPR responses to these variations are not conclusive and still need to be assessed.

Biodegradation, Environmental↗

Removal of phenolic compounds from a petrochemical effluent with a methanogenic consortium.

A methanogenic consortium was used to degrade phenol and ortho- (o-) cresol from a specific effluent of a petrochemical refinery. This effluent did not meet the local environmental regulations for phenolic compounds (178 mg/L), oils and greases (61 mg/L), ammoniacal nitrogen (75 mg/L) or sulfides (3.2 mg/L). The consortium, which degrades phenol via its carboxylation to benzoic acid, was progressively adapted to the effluent. Despite the very high effluent toxicity (EC50 of 2% with Microtox), the adapted consortium degraded 97% of 156 mg/L phenol in the supplemented effluent after 13 days in batch cultures (serum bottle). The addition of proteose peptone to the effluent is essential for phenol degradation. o-cresol was also transformed but not meta- or para-cresols. A continuous flow fixed-film anaerobic bioreactor was developed with the consortium. Treating the effluent with the bioreactor reduced phenol and phenolic compounds concentrations by 97 and 83%, respectively, for a hydraulic residence time of 6 h. This treatment also reduced by about half the effluent toxicity. Oils and greases and ammoniacal nitrogen were not affected. Similar microbiological forms were observed in serum bottles and in the bioreactors with or without the petrochemical effluent. These results indicate that this methanogenic consortium can treat efficiently the phenolic compounds in this specific petrochemical effluent.

Bacteria, Anaerobic↗

Environmental dissipation of fungicide triphenyltin acetate and its potential as a groundwater contaminant.

The effects of various factors on fungicide triphenyltin acetate dissipation in selected soil types were studied; temperature is clearly the most important factor of these. Residues of this fungicide in Fangtzupo clay loam and Lanyang loam incubated at 30 degrees C for 32 days (water content 100, 70, or 40% of field capacity) were 25% of the original amounts. In soil samples incubated at 10, 30, or 40 degrees C with 100% field capacity water content, residues were 80, 26, and 5.5% for Fangtzupo soil, respectively, and 83, 26, and 7.0% for Lanyang soil, respectively. No significant difference in dissipation rate was found between sterilized and unsterilized soil, suggesting that microbial degradation is not an important factor in this process. Rapid degradation rates were observed-half-lives of 8.3 to 19.4 days in Fangtzupo clay loam and 8.0 to 16.3 days in Lanyang loam at temperatures ranging from 30 to 40 degrees C (soil water content of 100, 70, and 40% of field capacity). Significant increases in half-life (approximately 150 days) occurred at lower temperatures (10 degrees C) in both soil types. The potential of triphenyltin acetate to contaminate groundwater was tested using the behavior assessment and groundwater pollution-potential models. Results indicate that the leaching rate of the fungicide is very slow (1.0 cm day(-1)) under normal conditions.

Biodegradation, Environmental↗

Biotreatability and kinetics of UASB reactor to mixtures of chlorophenol pollutants.

In most natural ecosystems heterotrophic microorganisms encounter complex mixtures of carbon sources, each of which is present at a concentration of few micrograms per litre. This study examined the biotreatability and kinetics of an upflow anaerobic sludge blanket (UASB) reactor to complex mixtures of chlorophenols encountered in environmental conditions using on-line and off-line experimental studies. Results indicate that (1) steady-state concentration was quite lower (98.3 mg L(-1)) with complex mixture of chlorophenols than steady-state concentration achieved when only 2.4 dichlorophenol (124 mg L(-1)) was studied alone on the same reactor; (2) that toxic effects of chlorophenols increase with increasing concentrations of toxicant. (Onset of the inhibitory effect occurred at a lower concentration in multi-substrate than in single substrate utilization): (3) addition of alternative utilizable substrate can mitigate toxic effects and enhance degradation; (4) the relative concentration of substrate was critical in determining utilization patterns. HPLC analysis of off-line experimental samples resulted in a steady-state treatment efficiency of 68% for COD, 36% for 2-chlorophenol, 40.5% for 4-chlorophenol, 70.7% for 2,4-dichlorophenol, 53.2% for 2,4,6-trichlorophenol and 42% for pentachlorophenol in presence of glucose. Kinetic constant in terms of Vmax and Ks were determined. Ks for the five chlorophenols ranged between 0.016 and 0.117 kg m(-3) day(-1) while Vmax range between 0.056 and 0.244 kg m(-3) day(-1).

Bacteria, Anaerobic↗

Phosphonate utilization by bacterial cultures and enrichments from environmental samples.

A selection of axenic microbial strains and a variety of environmental samples were investigated with respect to the utilization of a series of natural and xenobiotic phosphonates as the sole phosphorus source for growth. Phosphonate degradation was observed only with bacteria and not with eucaryotic microorganisms. All representatives of the phosphonates examined supported bacterial growth, with the exception of methylphosphonate diethylester. Yet, distinctly different phosphonate utilization patterns were noted between phosphonate-positive strains. C-P bond cleavage by a photosynthetic bacterium is reported for the first time; growing photoheterotrophically, Rhodobacter capsulatus ATCC 23782 was able to utilize 2-aminoethylphosphonate and alkylphosphonates. Bacteria with the potential to utilize at least one of the phosphonate moieties from the xenobiotic phosphonates Dequest 2010, Dequest 2041, and Dequest 2060 were detected in all environments, with only two exceptions for Dequest 2010. Phosphonate P utilization to an extent of 94 and 97%, for Dequest 2010 and Dequest 2041, respectively, provided evidence that a complete breakdown of these compounds with respect to the C-P bond cleavage can be achieved by some bacteria. The results suggest that phosphonate-utilizing bacteria are ubiquitous, and that selected strains can degrade phosphonates that are more complex than those described previously.

Bacteria↗

Transformation of nitrogen during vermicomposting of fly ash.

In view of environmental problems generated by large-scale production of fly ash, increasing attention is now being paid to the recycling fly ash as a source of plant nutrients in agriculture. However, the low amount of nitrogen in such materials forms a major constraint for such application. In the present study, the possibility of improving the N status in mixtures of fly ash and organic matter was investigated by adopting vermicomposting technology. Different combinations of fly ash and cow (Bos taurus) dung; namely fly ash alone, cow dung alone and fly ash + cow dung at 1:1, 1:3 and 3:1 ratios were incubated with and without epigeic earthworms (Eisenia fetida) for 50 days. The occurrence of N in different bio-available forms; namely easily mineralizable, NH4+ and NO3- tended to increase considerably in the series treated with earthworms. This behaviour was attributed primarily to increased microbiological activity in the vermicomposted samples and also to a considerable rise in the concentration of nitrogen-fixing bacteria in this series. Among the three combinations of vermicomposted fly ash and cow dung, the 1:1 mixture appeared to exhibit the highest availability of nitrogen.

Animals↗

Environmental assessment of 5-ethylidene-2-norbornene using modeled and measured fate and effects results.

Several predictive models were used to assess aquatic exposure, persistence (P) and potential for long-range transport (LRT) of 5-ethylidene-2-norbornene (ENB). Such estimations are components of the assessment process for persistent, bioaccumulative and toxic (PBT) substances, which are also referred to as persistent organic pollutants (POPs). An ecological exposure assessment for ENB from manufacturing activities was conducted based on physical/chemical properties, monitoring data, and degradation, transport and distribution estimates. Based on the results of several model predictions, chronic exposure of aquatic organisms is not expected, due to the anticipated residence time of ENB in aquatic ecosystems. These modeled results consistently predict ENB does not present the potential to persist in the environment. Volatilization from water to the air is calculated to occur at a relatively rapid rate for ENB based on its Henry's Law constant. Once in the air, ENB is expected to degrade rapidly due to oxidation by hydroxyl radicals and ozone based on calculated atmospheric half-lives of 57 and 27 min, respectively. Additionally, ENB is not predicted to undergo long-range transport based on the short atmospheric half-life due to oxidation by hydroxyl radicals and ozone. Additionally, based on predicted exposure from site-specific emission using the EPA model EFAST, ENB is not expected to reach concentrations of concern for chronic aquatic toxicity endpoints.

Air↗

Introduction of a de novo bioremediation ability, aryl reductive dechlorination, into anaerobic granular sludge by inoculation of sludge with Desulfomonile tiedjei.

Methanogenic upflow anaerobic granular-sludge blanket (UASB) reactors treat wastewaters at a high rate while simultaneously producing a useful product, methane; however, recalcitrant environmental pollutants may not be degraded. To impart 3-chlorobenzoate (3-CB)-dechlorinating ability to UASB reactors, we inoculated granular sludge in UASB reactors with either a pure culture of Desulfomonile tiedjei (a 3-CB-dechlorinating anaerobe) or a three-member consortium consisting of D. tiejei, a benzoate degrader, and an H2-utilizing methanogen. No degradation occurred in an uninoculated control reactor which was started with the same granular sludge, but inoculated reactors and granules from the inoculated UASB systems rapidly transformed 3-CB (54 mumol/day/g of granule biomass). After several months at a hydraulic retention time of 0.5 day, much shorter than the generation time of D. tiedjei, the reactors still dechlorinated 3-CB. This indicated that the bacteria were immobilized in the reactor granules, and by using an antibody probe for D. tiedjei, we demonstrated that this microorganism had colonized the sludge granules. These results represent the first addition of a pure culture or a defined microbial mixture to a viable waste treatment process to introduce a specific de novo degradative pathway into a granular-sludge consortium.

Anaerobiosis↗

Purification to homogeneity and characterization of a novel Pseudomonas putida chromate reductase.

Cr(VI) (chromate) is a widespread environmental contaminant. Bacterial chromate reductases can convert soluble and toxic chromate to the insoluble and less toxic Cr(III). Bioremediation can therefore be effective in removing chromate from the environment, especially if the bacterial propensity for such removal is enhanced by genetic and biochemical engineering. To clone the chromate reductase-encoding gene, we purified to homogeneity (>600-fold purification) and characterized a novel soluble chromate reductase from Pseudomonas putida, using ammonium sulfate precipitation (55 to 70%), anion-exchange chromatography (DEAE Sepharose CL-6B), chromatofocusing (Polybuffer exchanger 94), and gel filtration (Superose 12 HR 10/30). The enzyme activity was dependent on NADH or NADPH; the temperature and pH optima for chromate reduction were 80 degrees C and 5, respectively; and the K(m) was 374 microM, with a V(max) of 1.72 micromol/min/mg of protein. Sulfate inhibited the enzyme activity noncompetitively. The reductase activity remained virtually unaltered after 30 min of exposure to 50 degrees C; even exposure to higher temperatures did not immediately inactivate the enzyme. X-ray absorption near-edge-structure spectra showed quantitative conversion of chromate to Cr(III) during the enzyme reaction.

Biodegradation, Environmental↗

Biocomposites of plasticized starch reinforced with cellulose crystallites from cottonseed linter.

Environmentally friendly starch biocomposites were successfully developed using a colloidal suspension of cottonseed linter cellulose crystallite as a filler to reinforce glycerol plasticized starch (PS). The cellulose crystallites, having lengths of 350 +/- 70 nm and diameters of 40 +/- 8 nm on average, were prepared from cottonseed linters by acid hydrolysis. The dependence of morphology and properties of the PS-based biocomposites on cellulose crystallites content in the range from 0 to 30 wt.-% was investigated by scanning electron microscopy, differential scanning thermal analysis, dynamic mechanical thermal analysis, and measurements of mechanical properties and water absorption. The results indicate that the strong interactions between fillers and between the filler and PS matrix play a key role in reinforcing the resulting composites. The PS/cellulose crystallite composites, conditioned at 50% relative humidity, undergo an increase in both tensile strength and Young's modulus from 2.5 MPa for PS film to 7.8 MPa and from 36 MPa for PS film to 301 MPa. Further, incorporating cottonseed linter cellulose crystallites into PS matrix leads to an improvement in water resistance for the resulting biocomposites. The mechanical behaviors of the starch-based biocomposites as a function of cellulose crystallites content.

Biodegradation, Environmental↗

Distribution of the coenzyme M pathway of epoxide metabolism among ethene- and vinyl chloride-degrading Mycobacterium strains.

An epoxyalkane:coenzyme M (CoM) transferase (EaCoMT) enzyme was recently found to be active in the aerobic vinyl chloride (VC) and ethene assimilation pathways of Mycobacterium strain JS60. In the present study, EaCoMT activity and genes were investigated in 10 different mycobacteria isolated on VC or ethene from diverse environmental samples. In all cases, epoxyethane metabolism in cell extracts was dependent on CoM, with average specific activities of EaCoMT between 380 and 2,910 nmol/min/mg of protein. PCR with primers based on conserved regions of EaCoMT genes from Mycobacterium strain JS60 and the propene oxidizers Xanthobacter strain Py2 and Rhodococcus strain B-276 yielded fragments (834 bp) of EaCoMT genes from all of the VC- and ethene-assimilating isolates. The Mycobacterium EaCoMT genes form a distinct cluster and are more closely related to the EaCoMT of Rhodococcus strain B-276 than that of Xanthobacter strain Py2. The incongruence of the EaCoMT and 16S rRNA gene trees and the fact that isolates from geographically distant locations possessed almost identical EaCoMT genes suggest that lateral transfer of EaCoMT among the Mycobacterium strains has occurred. Pulsed-field gel electrophoresis revealed large linear plasmids (110 to 330 kb) in all of the VC-degrading strains. In Southern blotting experiments, the strain JS60 EaCoMT gene hybridized to many of the plasmids. The CoM-mediated pathway of epoxide metabolism appears to be universal in alkene-assimilating mycobacteria, possibly because of plasmid-mediated lateral gene transfer.

Biodegradation, Environmental↗

Environmental exposure assessment of fluoroquinolone antibacterial agents from sewage to soil.

The behavior of fluoroquinolone antibacterial agents (FQs) during mechanical-biological wastewater treatment was studied by mass flow analysis. In addition, the fate of FQs in agricultural soils after sludge application was investigated. Concentrations of FQs in filtered wastewater (raw sewage, primary, secondary, and tertiary effluents) were determined using solid-phase extraction with mixed phase cation exchange disk cartridges and reversed-phase liquid chromatography with fluorescence detection. FQs in suspended solids, sewage sludge (raw, excess, and anaerobically digested sludge), and sludge-treated soils were determined as described for the aqueous samples but preceded by accelerated solvent extraction. Wastewater treatment resulted in a reduction of the FQ mass flow of 88-92%, mainly due to sorption on sewage sludge. A sludge-wastewater partition coefficient (log Kd approximately 4) was calculated in the activated sludge reactors with a hydraulic residence time of about 8 h. No significant removal of FQs occurred under methanogenic conditions of the sludge digesters. These results suggest sewage sludge as the main reservoir of FQ residues and outline the importance of sludge management strategies to determine whether most of the human-excreted FQs enter the environment. Field experiments of sludge-application to agricultural land confirmed the long-term persistence of trace amounts of FQs in sludge-treated soils and indicated a limited mobility of FQs into the subsoil.

Adsorption↗

Substrate inhibition kinetics for toluene and benzene degrading pure cultures and a method for collection and analysis of respirometric data for strongly inhibited cultures.

We present an evaluation of the qualitative and quantitative effects that high concentrations of benzene and toluene have on the growth rate of several pure cultures that use these compounds as their sole carbon and energy source. The cultures employed were five widely studied environmental isolates: Pseudomonas putida F1, P. putida mt2, P. mendocina KR, Ralstonia pickettii PKO1, and Burkholderia cepacia G4. Three cultures degraded toluene following a pattern consistent with the kinetic model of Wayman and Tseng (1976) while the other two followed a modification of this model introduced by Alagappan and Cowan (2001). The pattern followed for benzene degradation was different than that for toluene degradation for all four capable pure cultures and consistent with that described by the model of Luong (1987). Mechanisms of substrate inhibition and solvent toxicity are discussed, used to conceptually evaluate the reasons for the differences in inhibition behavior, and used to support a call for more widespread use of the empirical, terminal substrate concentration inhibition models employed here. We also present the methodology developed to overcome a limitation commonly encountered when attempting to collect oxygen uptake data for use in quantifying substrate inhibition kinetics. The experimental method was effective for use in the collection of high quality data and the substrate inhibition models most useful in representing the growth of bacteria on these solvents are those that show a complete loss of activity at high concentration rather than the more popular asymptotic inhibition models.

Benzene↗

An overview on olive mill wastes and their valorisation methods.

Olive mill wastes represent an important environmental problem in Mediterranean areas where they are generated in huge quantities in short periods of time. Their high phenol, lipid and organic acid concentrations turn them into phytotoxic materials, but these wastes also contain valuable resources such as a large proportion of organic matter and a wide range of nutrients that could be recycled. In this article, recent research studies for the valorisation of olive mill wastes performed by several authors were reviewed: second oil extraction, combustion, gasification, anaerobic digestion, composting and solid fermentation are some of the methods proposed. Special attention was paid to the new solid waste generated during the extraction of olive oil by the two-phase system. The peculiar physicochemical properties of the new solid waste, called two-phase olive mill waste, caused specific management problems in the olive mills that have led to the adaptation and transformation of the traditional valorisation strategies. The selection of the most suitable or appropriate valorisation strategy will depend on the social, agricultural or industrial environment of the olive mill. Although some methods are strongly consolidated in this sector, other options, more respectful with the environment, should also be considered.

Biodegradation, Environmental↗