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Current and future applications of flow cytometry in aquatic microbiology.

Flow cytometry has become a valuable tool in aquatic and environmental microbiology that combines direct and rapid assays to determine numbers, cell size distribution and additional biochemical and physiological characteristics of individual cells, revealing the heterogeneity present in a population or community. Flow cytometry exhibits three unique technical properties of high potential to study the microbiology of aquatic systems: (i) its tremendous velocity to obtain and process data; (ii) the sorting capacity of some cytometers, which allows the transfer of specific populations or even single cells to a determined location, thus allowing further physical, chemical, biological or molecular analysis; and (iii) high-speed multiparametric data acquisition and multivariate data analysis. Flow cytometry is now commonly used in aquatic microbiology, although the application of cell sorting to microbial ecology and quantification of heterotrophic nanoflagellates and viruses is still under development. The recent development of laser scanning cytometry also provides a new way to further analyse sorted cells or cells recovered on filter membranes or slides. The main infrastructure limitations of flow cytometry are: cost, need for skilled and well-trained operators, and adequate refrigeration systems for high-powered lasers and cell sorters. The selection and obtaining of the optimal fluorochromes, control microorganisms and validations for a specific application may sometimes be difficult to accomplish.

Animals↗

A sustainable approach towards rural development: dry toilets in Nepal.

Existing inadequate sewerage systems and direct disposal of household waste into water courses has tremendously increased water pollution. Dry toilets are feasible in rural and peri-urban areas to reduce the consumption of costlier water that is required for flushing. As conventional treatment technologies require high investment, and operation and maintenance costs, dry toilets are the only suitable option left for sanitation in the 21st century when working with limited financial resources. To reduce environmental degradation and overcome this problem, the dry toilet is only the realistic option in Nepal. Two reactors, one exposed to sunrays and the other without sunrays, were constructed. In the model with sunrays, it was found that in 48 days of observation faecal coliform presence depleted to 610 cells per gm from the initial value of 7 x 10(10) and volatile organic matter came down from 98.09% to 70.18%. Similarly, in the other model, the destruction of faecal coliform in 65 days was found to be 920 cells/gm while the destruction of organic matter took 75 days. Also, observing from 313 people in a cluster on the pilot project, the annually recovered value of N, P and K was found to be 1565 kg, 125 kg, 344 kg, respectively. This paper deals with the different types of dry ecological toilet, their performance and feasibility study in Nepal, with the full involvement of local people, based on complete laboratory analysis and regular monitoring. Using dry toilets will save 14 LPCD, which is equivalent to 14 MLD and the resulting demand will become only 80 MLD for the urban area of Kathmandu. The result advocates the implementation of ecological dry toilets to save valuable water wasted in flushing, as well as saving the resources needed to treat the waste.

Biodegradation, Environmental↗

Partial characterization of the polluting load of swine wastewater treated with an integrated biodigestion system.

The stabilization of swine wastewaters from swine confined housing by the combination of a upflow anaerobic sludge blanket (UASB) reactor and waste stabilization ponds is a viable alternative to minimize the environmental impact caused by inadequate disposal of swine wastewaters. In the present study, the polluting load of pre-decanted swine wastewater treated with a series of two 0.705 m(3) UASB reactors and then in parallel in aerated and non-aerated stabilization tanks was investigated from January to July, 2000. Physicochemical and microbiological analyses were made adopting standard methods (Standard Methods for Examination of Water and Wastewater, 19th ed., American Public Health Association, Washington, DC, 1995). COD values decreased as the wastewater ran through the integrated biodigestion system dropping from about 3492+/-511-4094 mgl(-1)+/-481 to 124+/-52-490 mgl(-1)+/-230, while nitrate and nitrite levels increased in stabilization tanks, ranging respectively from 4+/-0 to 20 mgl(-1)+/-3 and 3+/-1 to 11 mgl(-1)+/-24. Although the removal of Escherichia coli was more than 97%+/-6, the effluents of the treatment system still contained unacceptable levels of E. coli (1.6 x 10(3)-1.2 x 10(6) 100 ml(-1)) according to WHO guidelines for use of wastewater in agriculture and aquaculture. These results indicate the necessity of changes on operational characteristics of the treatment system such as an increase of the hydraulic retention time in UASB reactors or in stabilization tanks.

Anaerobiosis↗

Considerations necessary in gathering occurrence data for selected unstable compounds in the USEPA Unregulated Contaminant Candidate List in USEPA Method 526.

U.S. Environmental Protection Agency (EPA) Method 526 was developed for the analysis of target analytes that are subject to degradation by hydrolysis. Two technical hurdles that had to be overcome were preservation of the target analytes and selection of a suitable solid-phase extraction material. The target analytes were diazinon, disulfoton, fonofos, terbufos, prometon, 1,2-diphenylhydrazine, nitrobenzene, acetochlor, 2,4,6-trichlorophenol, 2,4-dichlorophenol, and cyanazine. Diazolidinyl urea was used for the first time as a microbial inhibitor in an EPA drinking water method. Experiment confirmed antimicrobial agents containing copper or mercury salts increased hydrolysis degradation rates. Trisodium ethylenediaminetetraacetic acid salt was added to chelate metal ions that may increase hydrolysis rates. A pH 7 buffer of tris(hydroxymethyl)aminomethane (Tris) and Tris hydrochloride was used to minimize rates of hydrolysis. The use of ascorbic acid prevented degradation of 2,4-dichlorophenol, terbufos, fonofos, diazinon, and disulfoton due to residual chlorine. Samples were extracted using a styrene divinylbenzene solid-phase material and analyzed by capillary column gas chromatography/mass spectrometry. A 21-day storage stability study, together with precision and accuracy studies, showed that this method has suitable sensitivity, accuracy, precision, and ruggedness for use in the EPA's Unregulated Contaminant Monitoring Rule drinking water occurrence survey.

Biodegradation, Environmental↗

Subsurface irrigation as a microbial delivery tool for bioaugmentation: transport, distribution and survival in large packed soil columns.

Traditionally, soil surface inoculation and surface irrigation are used for delivery, transport and distribution of bacteria for agricultural and in-situ environmental decontamination applications. The objective of this study was to test whether subsurface irrigation with a water table management (WTM) system, successfully used previously for nutrient delivery, could also be used to deliver bacteria to soil depths. Twelve stainless steel columns, 1000 mm in length and 200 mm in diameter, were packed with a sandy loam soil. Four experimental treatments, two subsurface irrigation and two surface irrigation, were randomly allocated, in triplicate, to the twelve columns. The transport, implantation and survival of Sinorhizobium (Rhizobium) meliloti, strain A-025, in these columns were monitored at different depths and times after surface and subsurface irrigation. Even though the transport of bacterial cells was slower with the subsurface irrigation regime than with surface irrigation, more bacteria were implanted at different depths with subsurface irrigation. The numbers of bacterial cells distributed with subsurface irrigation were 1.6x10(5), 1.3x10(5), 2.6x10(5) and 2.9x10(5) cfu (colony forming units) x g(-1) of soil at 60, 300, 500 and 700 mm depths, respectively, whereas with surface irrigation the numbers were 2.0x10(4), 3.0x10(4), 1.9x10(5), 9.0x10(2) cfu x g(-1) of soil. These results dearly indicate that subirrigation can be used effectively to bioaugment a sandy loam soil matrix.

Agriculture↗

Biotransformation of Hg(II) by cyanobacteria.

The biotransformation of Hg(II) by cyanobacteria was investigated under aerobic and pH-controlled culture conditions. Mercury was supplied as HgCl(2) in amounts emulating those found under heavily impacted environmental conditions where bioremediation would be appropriate. The analytical procedures used to measure mercury within the culture solution, including that in the cyanobacterial cells, used reduction under both acid and alkaline conditions in the presence of SnCl(2). Acid reduction detected free Hg(II) ions and its complexes, whereas alkaline reduction revealed that meta-cinnabar (beta-HgS) constituted the major biotransformed and cellularly associated mercury pool. This was true for all investigated species of cyanobacteria: Limnothrix planctonica (Lemm.), Synechococcus leopoldiensis (Racib.) Komarek, and Phormidium limnetica (Lemm.). From the outset of mercury exposure, there was rapid synthesis of beta-HgS and Hg(0); however, the production rate for the latter decreased quickly. Inhibitory studies using dimethylfumarate and iodoacetamide to modify intra- and extracellular thiols, respectively, revealed that the former thiol pool was required for the conversion of Hg(II) into beta-HgS. In addition, increasing the temperature enhanced the amount of beta-HgS produced, with a concomitant decrease in Hg(0) volatilization. These findings suggest that in the environment, cyanobacteria at the air-water interface could act to convert substantial amounts of Hg(II) into beta-HgS. Furthermore, the efficiency of conversion into beta-HgS by cyanobacteria may lead to the development of applications in the bioremediation of mercury.

Biodegradation, Environmental↗

Dehalogenation of dichloroethene in a contaminated soil: fatty acids and alcohols as electron donors and an apparent requirement for tetrachloroethene.

Environmental soil contamination at an industrial site in Marion, Ohio (USA) with tetrachloroethene (perchloroethene, PCE) resulted in residual cis-1, 2-dichloroethene (DCE) contamination that had not declined after more than 15 years. Microcosm slurries containing 2.6% soil from this site were supplemented with different electron donors, i.e., individual fatty acids or alcohols. None of the microcosms supported complete DCE dechlorination, unless PCE was added to the microcosm at initiation. The addition of fresh PCE resulted in the dehalogenation of PCE to DCE in the microcosms supplemented with fatty acids having an even number of carbon atoms (acetate, butyrate, and caproate), but not in those with an odd number of carbon atoms (formate, propionate, and valerate), where negligible or no activity was detected. No significant further DCE degradation was observed in any of the microcosms supplied with fatty acids as electron donors. Microcosms supplemented with freshly added PCE bioconverted PCE to DCE and completely dehalogenated both the ex-novo and soil-supplied DCE within 60 days, but only if alcohols having an even number of carbon atoms (ethanol or butanol) were also added as electron donors. Odd-numbered alcohols either did not produce dehalogenation (as with methanol) or only dehalogenated PCE to DCE (as with propanol).

Alcohols↗

Fate of genetically-engineered bacteria in activated sludge microcosms.

The conclusions that can be derived from this study regarding the fate of the GEMs and their ability to degrade added pollutants are as follows: 1. Both GEMs were able to survive in the microcoms. Because Pseudomonas sp. B13 has been cultured for a long time in the laboratory, it was not expected to survive well in the microcosm. Surprisingly, it and the derivative GEMs persisted at a high population level of approximately 10(5) bacteria/ml. Pure culture studies had demonstrated an ability of FR1(pFRC20P) to readily degrade simultaneous mixtures of 3CB and 4MB. In the microcosms, however, the GEM did not perform as well as expected, particularly when confronted with a shock load of a 3CB and 4MB mixture. Thus, the microcosm studies may be of potential help for making predictions concerning environmental applications of GEMs. 2. Pseudomonas sp. B13 derivative strains FR1 and FR1(pFRC20P) were able to degrade low concentrations of substituted benzoates within the complex ecosystem of the activated sludge microcosm. A good deal of information concerning the degradation pathway for aromatics by Pseudomonas sp. B13 was already known. This allowed for the construction of the stable, regulated pathways for the degradation of substituted aromatic compounds in both GEMs and indicates that the construction of similar GEMs for the degradation of environmental pollutants is a promising experimental strategy. 3. There was not any demonstrable, adverse effect of GEM addition to the microbial population level in the microcosm. The GEMs were even able to function in a protective manner for the indigenous populations by buffering them against the adverse effects of addition of substituted benzoates. In contrast, for microcosms lacking GEM addition, a wash-out of the bacterial population in the microcosm occurred (data not shown). 4. Lateral transfer of new genetic information (xylXYZLS) from the GEM chromosome to indigenous microorganisms was not detected, whereas transfer of the hybrid, mobilizable pFRC20P carrying the gene for lactone isomerase did apparently occur. In this particular case, transfer may have been beneficial for the community as a whole if it increased the ecosystem's ability to cope with the presence of toxic pollutants. As more GEMs are constructed for specific biotechnological applications the diversity and complexity of microcosms used to study their fate and function will increase. The ability of such studies to predict a priori the fate of these microorganisms will help to develop strategies both to decrease the risks associated with introducing GEMs into the environment and to increase and regulate the capacity of GEMs to degrade toxic pollutants.(ABSTRACT TRUNCATED AT 400 WORDS)

Bacteria↗

Nitrite reduction by a mixed culture under conditions relevant to shortcut biological nitrogen removal.

Dissimilative reduction of nitrite by nitrite-acclimated cells was investigated in a batch reactor under various environmental conditions that can be encountered in shortcut biological nitrogen removal (SBNR: ammonia to nitrite and nitrite to nitrogen gas). The maximum specific nitrite reduction rate was as much as 4.3 times faster than the rate of nitrate reduction when individually tested, but the reaction was inhibited in the presence of nitrate when the initial nitrate concentration was greater than approximately 25 mg-N/1 or the initial NO3(-)N/NO2(-)-N ratio was larger than 0.5. Nitrite reduction was also inhibited by nitrite itself when the concentration was higher than that to which the cells had been acclimated. Therefore, it was desirable to avoid excessively high nitrite and nitrate concentrations in a denitrification reactor. Nitrite reduction, however, was not affected by an alkaline pH (in the range of 7-9) or a high concentration of FA (in the range of 16-39 mg/l), which can be common in SBNR processes. The chemical oxygen demand (COD) requirement for nitrite reduction was approximately 22-38% lower than that for nitrate reduction, demonstrating that the SBNR process can be economical. The specific consumption, measured as the ratio of COD consumed to nitrogen removed, was affected by the availability of COD and the physiological state of the cells. The ratio increased when the cells grew rapidly and were storing carbon and electrons.

Ammonia↗

Unlocking microbial potential: advances in omics and bioinformatics for aromatic hydrocarbon degradation.

Aromatic hydrocarbons (AHs) are persistent environmental pollutants with high toxicity. Bacterial degradation of AHs provides a sustainable and cost-effective approach for the remediation of sites contaminated with both mono- and polycyclic aromatic hydrocarbons. Aerobic degradation of AHs typically involves oxygenases-mediated hydroxylation followed by aromatic ring cleavage. In contrast, anaerobic degradation relies on diverse activation mechanisms that ultimately converge on the central intermediate benzoyl-CoA. Over the past decades, research on bacterial degradation of AHs has grown steadily, supported by advances in omics and bioinformatics. In this review, we summarize the current knowledge on the pathways, enzymes, and microbial diversity involved in AH degradation, highlighting how omics and bioinformatic approaches are advancing our understanding of this process. However, to improve our knowledge of microbial AHs catabolism, it is crucial to prioritize the characterization of novel enzymes and pathways, especially those mediating anaerobic and hybrid degradation strategies. Addressing this gap requires the development of specialized resources that incorporate a broader taxonomic diversity and an expanded inventory of anaerobic genes and enzymes supported by experimental evidence. Equally important is the integration of multi-omics technologies, artificial intelligence, and ecological modeling into unified analytical pipelines. These efforts will be key to fully unlocking microbial metabolic potential and guiding more effective bioremediation and monitoring strategies for AHs.

Biodegradation, Environmental↗

Transient response of aerobic and anoxic activated sludge activities to sudden substrate concentration changes.

The state-of-the-art understanding of activated sludge processes as summarized in activated sludge models (ASMs) predicts an instantaneous increase in the biomass activity (which is measured, e.g., by the corresponding respiration rate OUR, NUR, etc.) under sudden substrate concentration changes. Experimental data (e.g., short-term batch respiration experiments under aerobic or anoxic conditions) collected for the calibration of the dynamic models (ASMs) often exhibit a transient phenomenon while attaining maximum activity, which cannot be explained by the current understanding of the activated sludge process. That transient phenomenon exhibits itself immediately upon addition of a substrate source to an endogenously respiring activated sludge sample and it usually takes a few minutes until the activated sludge reaches its maximum possible rate under given environmental conditions. This discrepancy between the state-of-the-art model and the experimental data is addressed in detail in this investigation. It is shown that the discrepancy is not caused by an error in the experimental set-up/data but it is rather due to model inadequacy. Among the hypotheses proposed, it appears that this transient response of the activated sludge most likely results from the sequence of intracellular reactions involved in substrate degradation by the activated sludge. Results from studies performed elsewhere with pure cultures (S. cerevisae and E. coli) support the hypothesis. The transient phenomenon can be described by a dynamic metabolic network model or by a simple first-order model, as adopted in this study. The transient phenomenon occurring in short-term batch respiration experiments is shown to interfere severely with parameter estimation if not modeled properly (2.8%, 11.5%, and 16.8% relative errors [average of three experiments] on Y(H), micro(maxH), and K(S), respectively). Proper modeling of this transient phenomenon whose time constant is on the order of minutes (1 to 3 min) is expected to contribute fundamentally to a better understanding and modeling of Orbal, carousel, and SBR-type treatment plants with fast-alternating process conditions, although such studies are beyond the scope of this report.

Aerobiosis↗

Bioremediation of olive oil mill wastewater: chemical alterations induced by Azotobacter vinelandii.

An environmentally friendly bioremediation system of olive oil mill wastewater (OMWW) is studied with respect to its physicochemical characteristics and degradation efficiency on major characteristic constituents. The method exploits the biochemical versatility of the dinitrogen fixing bacterium Azotobacter vinelandii (strain A) to grow in OMWW at the expense of its constituents and to transform it into an organic liquid fertilizer. The system eliminates the phytotoxic principles from OMWW and concomitantly enriches it with an agriculturally beneficial microbial consortium along with useful metabolites of the latter. The end product, branded "biofertilizer", is used as soil conditioner and liquid organic fertilizer. Growth of A. vinelandii in OMWW results in the decline of content of most of the compounds associated with phytotoxicity, and this is confirmed by the assessment of degradation yields. In parallel, during the process several other compounds noncommittally undergo degradation and biotransformation. More specifically, the biofertilization system is capable of achieving removal yields as high as 90 and 96% after 3 and 7 days of treatment, respectively. Statistical analysis of the results showed that between the periods of operation no significant difference occurs with respect to the degradation yield. Moreover, the degradation yield from 3 to 7 days of continuous operation of the system remains almost unaltered during 2 consecutive years.

Azotobacter vinelandii↗

Ecological control analysis: being(s) in control of mass flux and metabolite concentrations in anaerobic degradation processes.

Identification of the functional groups of microorganisms that are predominantly in control of fluxes through, and concentrations in, microbial networks would benefit microbial ecology and environmental biotechnology: the properties of those controlling microorganisms could be studied or monitored specifically or their activity could be modulated in attempts to manipulate the behaviour of such networks. Herein we present ecological control analysis (ECA) as a versatile mathematical framework that allows for the quantification of the control of each functional group in a microbial network on its process rates and concentrations of intermediates. In contrast to current views, we show that rates of flow of matter are not always limited by a single functional group; rather flux control can be distributed over several groups. Also, control over intermediate concentrations is always shared. Because of indirect interactions, through other functional groups, the concentration of an intermediate can also be controlled by functional groups not producing or consuming it. Ecological control analysis is illustrated by a case study on the anaerobic degradation of organic matter, using experimental data obtained from the literature. During anaerobic degradation, fermenting microorganisms interact with terminal electron-accepting microorganisms (e.g. halorespirers, methanogens). The analysis indicates that flux control mainly resides with fermenting microorganisms, but can shift to the terminal electron-accepting microorganisms under less favourable redox conditions. Paradoxically, halorespiring microorganisms do not control the rate of perchloroethylene and trichloroethylene degradation even though they catalyse those processes themselves.

Bacteria, Anaerobic↗

Increasing bioavailability of phosphorus from fly ash through vermicomposting.

Due to the environmental problems created by large-scale fly ash generation throughout the world, efforts are being made to recycle these materials. An important component of the recycling effort is using fly ash to improve low-fertility soils. Because availability of many nutrients is very low in fly ash, available ranges of such nutrients must be improved to increase the effectiveness of fly ash as a soil amendment. In the present study, we assessed the possibility of increasing P bioavailability in fly ash through vermicomposting in a yard experiment. Fly ash was mixed with organic matter in the form of cow (Bos taurus) dung at 1:3, 1:1, and 3:1 ratios and incubated with and without epigeic earthworm (Eisenia fetida) for 50 d. The concentration of phosphate-solubilizing bacteria (PSB) was found to increase many fold in the earthworm-treated series of fly ash and organic matter combinations compared with the series without earthworm. This helped to transform considerable amounts of insoluble P from fly ash into more soluble forms and thus resulted in increased bioavailability of the nutrients in the vermicomposted series. Among different combinations of fly ash and organic matter, P availability in fly ash due to vermicomposting was significantly higher in the 1:1 fly ash to cow dung treatment compared with the other treatments.

Animals↗

Nitrous oxide emission and denitrification in chronically nitrate-loaded riparian buffer zones.

Riparian buffer zones are known to reduce diffuse N pollution of streams by removing and modifying N from agricultural runoff. Denitrification, often identified as the key N removal process, is also considered as a major source of the greenhouse gas nitrous oxide (N2O). The risks of high N2O emissions during nitrate mitigation and the environmental controls of emissions have been examined in relatively few riparian zones and the interactions between controls and emissions are still poorly understood. Our objectives were to assess the rates of N2O emission from riparian buffer zones that receive large loads of nitrate, and to evaluate various factors that are purported to control N emissions. Denitrification, nitrification, and N2O emissions were measured seasonally in grassland and forested buffer zones along first-order streams in The Netherlands. Lateral nitrate loading rates were high, up to 470 g N m(-2) yr(-1). Nitrogen process rates were determined using flux chamber measurements and incubation experiments. Nitrous oxide emissions were found to be significantly higher in the forested (20 kg N ha(-1) yr(-1)) compared with the grassland buffer zone (2-4 kg N ha(-1) yr(-1)), whereas denitrification rates were not significantly different. Higher rates of N2O emissions in the forested buffer zone were associated with higher nitrate concentrations in the ground water. We conclude that N transformation by nitrate-loaded buffer zones results in a significant increase of greenhouse gas emission. Considerable N2O fluxes measured in this study indicate that Intergovernmental Panel on Climate Change methodologies for quantifying indirect N2O emissions have to distinguish between agricultural uplands and riparian buffer zones in landscapes receiving large N inputs.

Agriculture↗

BTEX degradation by fungi.

Aromatic hydrocarbons such as benzene, toluene, ethylbenzene and xylenes (BTEX) are environmental pollutants that cause serious problems in groundwater, in surface water and in soil. Specifically, the coastal area of Bourgas bay (Bulgarian Black Sea coast) is very vulnerable to such pollution as the large petrochemical industry situated there poses a continuing threat to the environment. The potential for degradation of BTEX by fungi was studied. Two fungal strains were used in the experiments carried out in a batch reactor. The fungi were isolated from gasoline-polluted sites and were identified as deuteromycetes belonging to the Cladophialophora (CPH) and Cladosporium (CS) genera. It was found that neither of the fungal strains was able to degrade benzene. Ethylbenzene was easily degraded in all cases; o- and m-xylene were fully degraded by Cladophialophora sp. both as single substrates and in mixtures with toluene, whereas Cladosporium sp. was able to degrade them fully in mixtures with toluene only. p-Xylene was only partially degraded in all tests; however, the experiments with Cladosporium sp. showed better results compared to those with Cladophialophora sp. in this particular case. Ethylbenzene, o- and m-xylene negatively affected toluene removal rate of both fungal strains, which suggests possible competitive substrate interactions. In general, the results obtained with Cladophialophora sp. were better than those with Cladosporium sp., except in the case of p-xylene degradation.

Ascomycota↗

Effects of various environmental conditions on the transformation of chlorinated solvents by Methanosarcina thermophila cell exudates.

Several microbiologically produced biomolecules have been shown to degrade chlorinated contaminants found in groundwater systems. It was discovered that the cell-free exudates of the methanogen Methanosarcina thermophila were capable of carbon tetrachloride (CT) and chloroform (CF) degradation. Characterization of the exudates suggested that the active agents were porphorinogen-type molecules, possibly containing zinc. This research was performed to determine if the exudates from M. thermophila could be used for remediation purposes. The cell exudates were found to be capable of degrading CT, CF, tetrachloroethene, trichloroethene, and 1,1,1-trichloroethane. CT degradation was used to gauge exudate activity under a variety of conditions that would be encountered in the environment. The cell exudates were active when incubated in two types of soil matrices and at temperatures ranging from 4 to 23 degrees C. Over a 35-day period approximately 10.2 micromoles of CT were degraded by M. thermophila exudates. To test the hypothesis that the exudates contained either a zinc porphorinogen or a quinone, experiments were performed with zinc 5,10,15,20-tetra (4-pyridyl)-21 H, 23 H-porphine tetrakis, protoporphyrin IX zinc, and juglone. The two zinc porphyrins were capable of mediating CT degradation at rates comparable to those observed with the M. thermophila exudates; however, juglone was only capable of very slow CT transformation. The electron-transfer activity of the M. thermophila cell exudates was therefore more consistent with the activity of porphorinogens rather than quinones. Finally, in two enrichment cultures established from aquifer material and marine sediment, the possibility of excreted agents capable of degrading CT was evident.

Biodegradation, Environmental↗

Assessing and managing biological risks of plants used for bioremediation, including risks of transgene flow.

The plants used for phytoremediation pose special biological risks, whether transgenic or not, as most of the species: (a) are semi-domesticated; (b) are introduced from other habitats; (c) can become established in the contaminated site; (d) can spread and displace native species, and/or; (e) may introgress transgenes into related species. The addition of transgenes can reduce the risks, e.g. to sterilize or render the species and hybrid offspring hypersensitive to environmental effects (heat, cold), or to a chemical that will cull the species. Various measures can contain transgenes used in phytoremediation species to prevent gene flow, but most containment technologies are both uni-directional (prevent either outflow or influx), and are inherently leaky, even a concept specifically utilizable for phytoremediation--grafting non-transgenic scions on bioremediating transgenic rootstocks. Containment mechanisms should be either stacked with each other or with "mitigator" genes. Transgenic mitigation (TM) has mitigator genes added in tandem to the desired primary transgene (genetically linked) and the mitigator genes confer traits that are positive or neutral to the desired species but are deleterious to hybrids, keeping them at very low frequencies. The concept was demonstrated in tobacco and oilseed rape with a dwarfing mitigator gene that enhanced the reproductive productivity (harvest index) when cultured alone, but eliminated it from mixed populations. Besides the mitigator genes previously proposed for crop species (sterility, no seed shattering, dwarfing, no secondary dormancy) there are genes especially appropriate for phytoremediation, e.g. overexpression of cytokinin oxidase (reduces cytokinin levels) conferring reduced shoot systems (unfitness to compete) with a more extensive root system that is better for extracting toxic wastes as well as no-flowering for vegetatively propagated species. Thus, biotechnology can be harnessed to reduce risks from both non-transgenic and transgenic phytoremediation species.

Biodegradation, Environmental↗