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An organofluorophosphate-hydrolyzing activity in Tetrahymena thermophila.

An enzymatic activity that hydrolyzes O,O-diisoproplyphosphofluoridate (DFP) and O-1,2,2-trimethylpropylmethylphosphonofluoridate (Soman) was discovered in the ciliate protozoan Tetrahymena thermophila. The enzymatic activity classifies the protein as Mazur-type similar to that found in hog kidney and Escherichia coli. The rate of hydrolysis of Soman by the Tetrahymena-extract is the highest, on a per gram of extract basis, of any eucaryote. The molecular weight is approximately 75,400 as determined by Sephacryl column chromatography. A maximum fifteen-fold purification has been achieved. Potential exists for the detoxification and one-step detection of common organofluorophosphate pollutants. Additionally, Tetrahymena should prove an easier subject for manipulation than mammalian or squid sources. Protozoa may be a potentially important source of detoxification and degradation enzymes for other environmental contaminants.

Animals↗

Enrichment of mixed cultures capable of aerobic degradation of 1,2-dibromoethane.

1,2-dibromoethane (DBE) is a common environmental contaminant; it is potentially carcinogenic and has been detected in soil and groundwater supplies. Most of the biodegradation studies to date have been performed under anaerobic conditions or in the context of soil remediation, where the pollutant concentration was in the parts per billion range. In this work a mixed bacterial culture capable of complete aerobic mineralization of concentrations of DBE up to 1 g liter(-1) under well-controlled laboratory conditions was enriched. In order to verify biodegradation, formation of biodegradation products as well as the disappearance of DBE from the biological medium were measured. Complete mineralization was verified by measuring stoichiometric release of the biodegradation products. This mixed culture was found to be capable of degrading other halogenated compounds, including bromoethanol, the degradation of which has not been reported previously.

Aerobiosis↗

Bacterial dehalogenases: biochemistry, genetics, and biotechnological applications.

This review is a survey of bacterial dehalogenases that catalyze the cleavage of halogen substituents from haloaromatics, haloalkanes, haloalcohols, and haloalkanoic acids. Concerning the enzymatic cleavage of the carbon-halogen bond, seven mechanisms of dehalogenation are known, namely, reductive, oxygenolytic, hydrolytic, and thiolytic dehalogenation; intramolecular nucleophilic displacement; dehydrohalogenation; and hydration. Spontaneous dehalogenation reactions may occur as a result of chemical decomposition of unstable primary products of an unassociated enzyme reaction, and fortuitous dehalogenation can result from the action of broad-specificity enzymes converting halogenated analogs of their natural substrate. Reductive dehalogenation either is catalyzed by a specific dehalogenase or may be mediated by free or enzyme-bound transition metal cofactors (porphyrins, corrins). Desulfomonile tiedjei DCB-1 couples energy conservation to a reductive dechlorination reaction. The biochemistry and genetics of oxygenolytic and hydrolytic haloaromatic dehalogenases are discussed. Concerning the haloalkanes, oxygenases, glutathione S-transferases, halidohydrolases, and dehydrohalogenases are involved in the dehalogenation of different haloalkane compounds. The epoxide-forming halohydrin hydrogen halide lyases form a distinct class of dehalogenases. The dehalogenation of alpha-halosubstituted alkanoic acids is catalyzed by halidohydrolases, which, according to their substrate and inhibitor specificity and mode of product formation, are placed into distinct mechanistic groups. beta-Halosubstituted alkanoic acids are dehalogenated by halidohydrolases acting on the coenzyme A ester of the beta-haloalkanoic acid. Microbial systems offer a versatile potential for biotechnological applications. Because of their enantiomer selectivity, some dehalogenases are used as industrial biocatalysts for the synthesis of chiral compounds. The application of dehalogenases or bacterial strains in environmental protection technologies is discussed in detail.

Bacteria↗

Methanotrophic bacteria.

Methane-utilizing bacteria (methanotrophs) are a diverse group of gram-negative bacteria that are related to other members of the Proteobacteria. These bacteria are classified into three groups based on the pathways used for assimilation of formaldehyde, the major source of cell carbon, and other physiological and morphological features. The type I and type X methanotrophs are found within the gamma subdivision of the Proteobacteria and employ the ribulose monophosphate pathway for formaldehyde assimilation, whereas type II methanotrophs, which employ the serine pathway for formaldehyde assimilation, form a coherent cluster within the beta subdivision of the Proteobacteria. Methanotrophic bacteria are ubiquitous. The growth of type II bacteria appears to be favored in environments that contain relatively high levels of methane, low levels of dissolved oxygen, and limiting concentrations of combined nitrogen and/or copper. Type I methanotrophs appear to be dominant in environments in which methane is limiting and combined nitrogen and copper levels are relatively high. These bacteria serve as biofilters for the oxidation of methane produced in anaerobic environments, and when oxygen is present in soils, atmospheric methane is oxidized. Their activities in nature are greatly influenced by agricultural practices and other human activities. Recent evidence indicates that naturally occurring, uncultured methanotrophs represent new genera. Methanotrophs that are capable of oxidizing methane at atmospheric levels exhibit methane oxidation kinetics different from those of methanotrophs available in pure cultures. A limited number of methanotrophs have the genetic capacity to synthesize a soluble methane monooxygenase which catalyzes the rapid oxidation of environmental pollutants including trichloroethylene.

Base Sequence↗

Screening for exopolysaccharide-producing bacteria from sub-tropical polluted groundwater.

A selection of exopolysaccharide (EPS)--producing bacterial strains was conducted in groundwater adjacent to an old controlled landfill in the City of São Carlos (São Paulo, Brazil). The strains were isolated in P and E media under aerobic and microacrophilic conditions at 25 degrees C. A total of 26 strains were isolated and based on the mucoid mode of the colonies, 6 were selected and their morphological, physiological and biochemical aspects were characterized. All strains presented pigmentation, ranging from yellow to orange and from pink to salmon, with a shiny glistening aspect in all tested media. Strains Lb, Lc and Lg, which excelled the others with regard to the mucoid mode of the colonies, were selected to be cultured in E medium with alternate sucrose and glucose as carbon sources in anaerobiosis at 25 degrees C to analyze the production of EPS. Strains Lc and Lg were classified as being of order Actinomycelates, suborder Corynebacterineae. Lg strain was identified as Gordonia polyisoprenivorans and Lc strain did not correspond to a known description and therefore a more detailed study is under preparation. Considering all ecological aspects and the metabolic potential associated with the microorganisms of the environment studied, as well as the capacity to produce pigment and EPS, and the presence of G. polyisoprenivorans, a rubber degrader bacterium, the potential of the groundwater analyzed is evident as a source of microorganisms to be utilized in studies related to environmental remediation.

Actinomycetales↗

Small scale sanitation technologies.

Small scale systems can improve the sustainability of sanitation systems as they more easily close the water and nutrient loops. They also provide alternate solutions to centrally managed large scale infrastructures. Appropriate sanitation provision can improve the lives of people with inadequate sanitation through health benefits, reuse products as well as reduce ecological impacts. In the literature there seems to be no compilation of a wide range of available onsite sanitation systems around the world that encompasses black and greywater treatment plus stand-alone dry and urine separation toilet systems. Seventy technologies have been identified and classified according to the different waste source streams. Sub-classification based on major treatment methods included aerobic digestion, composting and vermicomposting, anaerobic digestion, sand/soil/peat filtration and constructed wetlands. Potential users or suppliers of sanitation systems can choose from wide range of technologies available and examine the different treatment principles used in the technologies. Sanitation systems need to be selected according to the local social, economic and environmental conditions and should aim to be sustainable.

Biodegradation, Environmental↗

Microbial degradation of polyurethane, polyester polyurethanes and polyether polyurethanes.

Polyurethane (PUR) is a polymer derived from the condensation of polyisocyanate and polyol and it is widely used as a base material in various industries. PUR, in particular, polyester PUR, is known to be vulnerable to microbial attack. Recently, environmental pollution by plastic wastes has become a serious issue and polyester PUR had attracted attention because of its biodegradability. There are many reports on the degradation of polyester PUR by microorganisms, especially by fungi. Microbial degradation of polyester PUR is thought to be mainly due to the hydrolysis of ester bonds by esterases. Recently, polyester-PUR-degrading enzymes have been purified and their characteristics reported. Among them, a solid-polyester-PUR-degrading enzyme (PUR esterase) derived from Comamonas acidovorans TB-35 had unique characteristics. This enzyme has a hydrophobic PUR-surface-binding domain and a catalytic domain, and the surface-binding domain was considered as being essential for PUR degradation. This hydrophobic surface-binding domain is also observed in other solid-polyester-degrading enzymes such as poly(hydroxyalkanoate) (PHA) depolymerases. There was no significant homology between the amino acid sequence of PUR esterase and that of PHA depolymerases, except in the hydrophobic surface-binding region. Thus, PUR esterase and PHA depolymerase are probably different in terms of their evolutionary origin and it is possible that PUR esterases come to be classified as a new solid-polyester-degrading enzyme family.

Bacteria↗

Comparison of EDTA and EDDS as potential soil amendments for enhanced phytoextraction of heavy metals.

Phytoextraction has been proposed as an alternative remediation technology for soils polluted with heavy metals or radionuclides, but is generally conceived as too slow working. Enhancing the accumulation of trace pollutants in harvestable plant tissues is a prerequisite for the technology to be practically applicable. The chelating aminopolycarboxylic acid, ethylene diamine tetraacetate (EDTA), has been found to enhance shoot accumulation of heavy metals. However, the use of EDTA in phytoextraction may not be suitable due to its high environmental persistence, which may lead to groundwater contamination. This paper aims to assess whether ethylene diamine disuccinate (EDDS), a biodegradable chelator, can be used for enhanced phytoextraction purposes. A laboratory experiment was conducted to examine mobilisation of Cd, Cu, Cr, Ni, Pb and Zn into the soil solution upon application of EDTA or EDDS. The longevity of the induced mobilisation was monitored for a period of 40 days after application. Estimated effect half lives ranged between 3.8 and 7.5 days for EDDS, depending on the applied dose. The minimum observed effect half life of EDTA was 36 days, while for the highest applied dose no decrease was observed throughout the 40 day period of the mobilisation experiment. Performance of EDTA and EDDS for phytoextraction was evaluated by application to Helianthus annuus. Two other potential chelators, known for their biodegradability in comparison to EDTA, were tested in the plant experiment: nitrilo acetic acid (NTA) and citric acid. Uptake of heavy metals was higher in EDDS-treated pots than in EDTA-treated pots. The effects were still considered insufficiently high to consider efficient remediation. This may be partly due to the choice of timing for application of the soil amendment. Fixing the time of application at an earlier point before harvest may yield better results. NTA and citric acid induced no significant effects on heavy metal uptake.

Biodegradation, Environmental↗

Oxidation of polychlorinated benzenes by genetically engineered CYP101 (cytochrome P450(cam)).

Polychlorinated benzenes are recalcitrant environmental pollutants primarily because they are resistant to attack by dioxygenases commonly used by micro-organisms for the biodegradation of aromatic compounds. We have investigated the oxidation of polychlorinated benzenes by mutants of the haem mono-oxygenase CYP101 (cytochrome P450(cam)) from Pseudomonas putida with the aim of generating novel systems for their biodegradation. Wild-type CYP101 had low activity for the oxidation of dichlorobenzenes and trichlorobenzenes to the chlorophenols, but no products were detected for the heavily chlorinated benzenes. Increasing the active-site hydrophobicity with the Y96F mutation increased the activity up to 100-fold, and both pentachlorobenzene and hexachlorobenzene were oxidized slowly to pentachlorophenol. Decreasing the space available at the top of the active site with the F87W mutation to force the substrate to be bound closer to the haem resulted in a further 10-fold increase in activity with most substrates. Introducing the F98W mutation, also at the top of the active site, decreased the NADH-turnover rates but increased the coupling efficiencies, and > 90% coupling was observed for 1,3-dichlorobenzene and 1,3,5-trichlorobenzene with the F87W--Y96F--F98W mutant. The V247L mutation generally increased the NADH-turnover rates, and the F87W--Y96F--V247L mutant showed reasonably fast NADH turnover (229 min(-1)) with the highly insoluble pentachlorobenzene without the need for surfactants or organic cosolvents. As all chlorophenols are degraded by micro-organisms, novel biodegradation systems could be constructed in which CYP101 mutants convert the inert polychlorinated benzenes to the phenols, which are then readily degraded by natural pathways.

Benzene Derivatives↗

Assessment of bioavailability of soil-sorbed atrazine.

Bioavailability of pesticides sorbed to soils is an important determinant of their environmental fate and impact. Mineralization of sorbed atrazine was studied in soil and clay slurries, and a desorption-biodegradation-mineralization (DBM) model was developed to quantitatively evaluate the bioavailability of sorbed atrazine. Three atrazine-degrading bacteria that utilized atrazine as a sole N source (Pseudomonas sp. strain ADP, Agrobacterium radiobacter strain J14a, and Ralstonia sp. strain M91-3) were used in the bioavailability assays. Assays involved establishing sorption equilibrium in sterile soil slurries, inoculating the system with organisms, and measuring the CO(2) production over time. Sorption and desorption isotherm analyses were performed to evaluate distribution coefficients and desorption parameters, which consisted of three desorption site fractions and desorption rate coefficients. Atrazine sorption isotherms were linear for mineral and organic soils but displayed some nonlinearity for K-saturated montmorillonite. The desorption profiles were well described by the three-site desorption model. In many instances, the mineralization of atrazine was accurately predicted by the DBM model, which accounts for the extents and rates of sorption/desorption processes and assumes biodegradation of liquid-phase, but not sorbed, atrazine. However, for the Houghton muck soil, which manifested the highest sorbed atrazine concentrations, enhanced mineralization rates, i.e., greater than those expected on the basis of aqueous-phase atrazine concentration, were observed. Even the assumption of instantaneous desorption could not account for the elevated rates. A plausible explanation for enhanced bioavailability is that bacteria access the localized regions where atrazine is sorbed and that the concentrations found support higher mineralization rates than predicted on the basis of aqueous-phase concentrations. Characteristics of high sorbed-phase concentration, chemotaxis, and attachment of cells to soil particles seem to contribute to the bioavailability of soil-sorbed atrazine.

Atrazine↗

Perchlorate as an environmental contaminant.

Perchlorate anion (ClO4-) has been found in drinking water supplies throughout the southwestern United States. It is primarily associated with releases of ammonium perchlorate by defense contractors, military operations, and aerospace programs. Ammonium perchlorate is used as a solid oxidant in missile and rocket propulsion systems. Traces of perchlorate are found in Chile saltpeter, but the use of such fertilizer has not been associated with large scale contamination. Although it is a strong oxidant, perchlorate anion is very persistent in the environment due to the high activation energy associated with its reduction. At high enough concentrations, perchlorate can affect thyroid gland functions, where it is mistakenly taken up in place of iodide. A safe daily exposure has not yet been set, but is expected to be released in 2002. Perchlorate is measured in environmental samples primarily by ion chromatography. It can be removed by anion exchange or membrane filtration. It is destroyed by some biological and chemical processes. The environmental occurrence, toxicity, analytical chemistry, and remediative approaches are discussed.

Biodegradation, Environmental↗

Characterization of the novel HCH-degrading strain, Microbacterium sp. ITRC1.

A gram-positive Microbacterium sp. strain, ITRC1, that was able to degrade the persistent and toxic hexachlorocyclohexane (HCH) isomers was isolated and characterized. The ITRC1 strain has the capacity to degrade all four major isomers of HCH present in both liquid cultures and aged contaminated soil. DNA fragments corresponding to the two initial genes involved in gamma-HCH degradative pathway, encoding enzymes for gamma-pentachlorocyclohexene hydrolytic dehalogenase (linB) and a 2,5-dichloro-2,5-cyclohexadiene-1,4-diol dehydrogenase (linC), were amplified by PCR and sequenced. Their presence in the ITRC1 genomic DNA was also confirmed by Southern hybridization. Sequencing of the amplified DNA fragment revealed that the two genes present in the ITRC1 strain were homologous to those present in Sphingomonas paucimobilis UT26. Both 16S rRNA sequencing and phylogenetic analysis resulted in the identification of the bacteria as a Microbacterium sp. We assume that these HCH-degrading bacteria evolved independently but possessed genes similar to S. paucimobilis UT26. The reported results indicate that catabolic genes for gamma-HCH degradation are highly conserved in diverse genera of bacteria, including the gram-positive groups, occurring in various environmental conditions.

Actinomycetales↗

Hexavalent chromium reduction by a dichromate-resistant gram-positive bacterium isolated from effluents of tanneries.

A gram-positive, chromium (Cr)-resistant bacterial strain (ATCC 700729) was isolated from effluent of tanneries. It was grown in media containing potassium dichromate concentration up to 80 mg ml(-1) of the medium. The dichromate reducing capability of the bacterium was checked by estimating the amount of Cr VI in the medium before and after introduction of bacterial culture. The influence of factors like pH of the medium, concentration of Cr, and the amount of the inoculum was studied to determine the ability of the bacterium to reduce Cr VI in the medium under various conditions. In a medium containing dichromate 20 mg ml(-1) more than 87% reduction of dichromate ions was achieved within 72 h. The feasibility of the use of this bacterial strain for detoxification of dichromate in the industrial wastewater has been assessed. The isolated strain can be exploited for specific environmental clean-up operations.

Biodegradation, Environmental↗

Dissimilatory Fe(III) and Mn(IV) reduction.

Dissimilatory Fe(III) and Mn(IV) reduction has an important influence on the geochemistry of modern environments, and Fe(III)-reducing microorganisms, most notably those in the Geobacteraceae family, can play an important role in the bioremediation of subsurface environments contaminated with organic or metal contaminants. Microorganisms with the capacity to conserve energy from Fe(III) and Mn(IV) reduction are phylogenetically dispersed throughout the Bacteria and Archaea. The ability to oxidize hydrogen with the reduction of Fe(III) is a highly conserved characteristic of hyperthermophilic microorganisms and one Fe(III)-reducing Archaea grows at the highest temperature yet recorded for any organism. Fe(III)- and Mn(IV)-reducing microorganisms have the ability to oxidize a wide variety of organic compounds, often completely to carbon dioxide. Typical alternative electron acceptors for Fe(III) reducers include oxygen, nitrate, U(VI) and electrodes. Unlike other commonly considered electron acceptors, Fe(III) and Mn(IV) oxides, the most prevalent form of Fe(III) and Mn(IV) in most environments, are insoluble. Thus, Fe(III)- and Mn(IV)-reducing microorganisms face the dilemma of how to transfer electrons derived from central metabolism onto an insoluble, extracellular electron acceptor. Although microbiological and geochemical evidence suggests that Fe(III) reduction may have been the first form of microbial respiration, the capacity for Fe(III) reduction appears to have evolved several times as phylogenetically distinct Fe(III) reducers have different mechanisms for Fe(III) reduction. Geobacter species, which are representative of the family of Fe(III) reducers that predominate in a wide diversity of sedimentary environments, require direct contact with Fe(III) oxides in order to reduce them. In contrast, Shewanella and Geothrix species produce chelators that solubilize Fe(III) and release electron-shuttling compounds that transfer electrons from the cell surface to the surface of Fe(III) oxides not in direct contact with the cells. Electron transfer from the inner membrane to the outer membrane in Geobacter and Shewanella species appears to involve an electron transport chain of inner-membrane, periplasmic, and outer-membrane c-type cytochromes, but the cytochromes involved in these processes in the two organisms are different. In addition, Geobacter species specifically express flagella and pili during growth on Fe(III) and Mn(IV) oxides and are chemotactic to Fe(II) and Mn(II), which may lead Geobacter species to the oxides under anoxic conditions. The physiological characteristics of Geobacter species appear to explain why they have consistently been found to be the predominant Fe(III)- and Mn(IV)-reducing microorganisms in a variety of sedimentary environments. In comparison with other respiratory processes, the study of Fe(III) and Mn(IV) reduction is in its infancy, but genome-enabled approaches are rapidly advancing our understanding of this environmentally significant physiology.

Archaea↗

A feasibility study of a Salix viminalis gravel hydroponic system to renovate primary settled wastewater.

A Salix viminalis/gravel system based on hydroponics was developed for wastewater renovation in order to avoid the problems of soil damage and pollution associated with long-term application of wastewater to soil. For such a system to work the mineral elements applied must match closely the requirements of the tree species. To examine this the growth and nutrient uptake of S. viminalis in wastewater was compared with that in Long Ashton nutrient solution (1/4 strength). S. viminalis grew more slowly in wastewater than in Long Ashton solution, but exhibited no obvious deficiency or toxicity symptoms. Since industrial wastewaters often contain metals, the extent to which copper might inhibit wastewater treatment in this system was also examined. S. viminalis was grown in wastewater amended with 10 and 100 ppm copper. Trees were unaffected by wastewater with 10 ppm copper when compared to trees grown in wastewater alone. Wastewater containing with 100 ppm copper was too toxic for the trees to thrive and wastewater treatment was reduced. Treatment efficiencies for unamended wastewater were 57.7% for nitrogen, 90.6% for phosphorus and 24.9% for potassium. These efficiencies are much greater than those quoted for a Salix/soil system, and thus Salix/gravel systems may have potential for wastewater treatment in environmentally sensitive areas or situations.

Biodegradation, Environmental↗

Fractionation of platinum group elements in aqueous systems: comparative kinetics of palladium and platinum removal from seawater by Ulva lactuca L.

A marine macroalga, Ulva lactuca L., was used as a substrate to compare the kinetics of palladium (Pd) and platinum (Pt) removal from seawater. This work indicates that, while the equilibrium behaviors of Pd and Pt are in many respects similar, their kinetic behaviors are quite distinct. The removal of both Pt(II) and Pt(IV) from seawater by U. lactuca is slower than the removal of Pd(II) by approximately an order of magnitude. Relative Pd and Pt removal rates are strongly influenced by system hydrodynamics. Under quiescent conditions, lambda(Pd)/lambda(Pt), the ratio of Pd and Pt removal rates, is 7 +/- 2, whereas under turbulent conditions lambda(Pd)/lambda(Pt) can be as large as 27. These observations suggest that the disparate kinetic behaviors of Pd and Pt may produce considerable differences in the environmental dispersion of these elements.

Biodegradation, Environmental↗

Biotechnological possibilities for waste tyre-rubber treatment.

Every year large amounts of spent rubber material, mainly from rubber tyres, are discarded. Of the annual total global production of rubber material, which amounts to 16-17 million tonnes, approximately 65% is used for the production of tyres. About 250 millions spent car tyres are generated yearly in USA only. This huge amount of waste rubber material is an environmental problem of great concern. Various ways to remediate the problem have been proposed. Among these are road fillings and combustion in kilns. Spent tyres, however, comprise valuable material that could be recycled if a proper technique can be developed. One way of recycling old tyres is to blend ground spent rubber with virgin material followed by vulcanization. The main obstacle to this recycling is bad adhesion between the crumb and matrix of virgin rubber material due to little formation of interfacial sulphur crosslinks. Micro-organisms able to break sulphur-sulphur and sulphur-carbon bonds can be used to devulcanize waste rubber in order to make polymer chains on the surface more flexible and facilitate increased binding upon vulcanization. Several species belonging to both Bacteria and Archaea have this ability. Mainly sulphur oxidizing species, such as different species of the genus Thiobacillus and thermoacidophiles of the order of Sulfolobales, have been studied in this context. The present paper will give a background to the problem and an overview of the biotechnological possibilities for solutions of waste rubber as an environmental problem, focusing on microbial desulphurization.

Bacteria↗