As pollution becomes an ever-increasing threat to the global environment pressure is being placed upon industry to "clean-up" its act, both in terms of reducing the possibility of new pollution and cleaning up already contaminated areas. It was with this in mind that Amplats embarked on a phytoremediation project to decontaminate nickel-polluted soils at one of their mine sites in Rustenburg, using the nickel hyperaccumulating plant, Berkheya coddii, which is endemic to the serpentine areas near Barberton, Mpumalanga. Besides the applied aspects pertaining to the development of the phytoremediation process we were also interested in more academic aspects concerning the transport and storage of nickel within the plant tissues. In order that the progress of nickel could be followed through the plant, a radio-tracer of ⁶³nickel was placed in the soil and its movement within the plant followed by analysing the plant material, at set intervals, using a liquid scintillation counter. From these studies it was found that the nickel appeared to be transported from the roots to the leaves of the plant via the xylem. It appeared that the nickel was not confined to the leaf to which it was initially transported and so movement of nickel within the phloem also appears to occur in B. coddii. As nickel is generally toxic to most plants, hyperaccumulators contain elements that nullify the toxic effect of nickel. In the case of Berkheya coddii it is thought that the accumulated nickel is bound to malate to form a harmless nickel complex. With this in mind an assay for L-malic acid was developed in order that any effect on L-malic acid, caused by growing Berkheya coddii on soils containing various concentrations of nickel, could be determined. This method also enabled comparisons of L-malic acid concentrations to be made between hyperaccumulators and non-hyperaccumulators of various plant species. From the L-malic acid comparisons it was found that the nickel concentration within soils affected the levels of L-malic acid within B. coddii and that the levels of L-malic acid within B. coddii were greater than that of a closely related non-hyperaccumulator, suggesting that L-malic acid is indeed involved in the hyperaccumulation mechanism within B. coddii. B. coddii was chosen as the tool in nickel phytoremediation at Rustenburg Base Metal Refineries as it was found to accumulate up to 2.5% nickel in the dry biomass, it grows rapidly and has a large above-ground biomass with a well developed root system, and it is perennial and so does not need to be planted each season. Earlier work had shown that the nickel levels in the roots were comparatively low (up to 0.3% nickel in the dry material) and thus, for ease of harvesting and to ensure the continued vegetative growth of the plant on the planted sites, it was decided that the leaves and stems of the plants would be harvested at the end of each growing season. The plant was also found to accumulate low levels (0.006 - 0.3 %) of precious metals, including platinum, palladium and rhodium, within its above ground biomass, making it attractive for the remediation of certain soils that contain low levels of these metals. Before B. coddii could be introduced to the Rustenburg area a comparison of the climatic and soil conditions of Barberton, the area to which B. coddii is endemic, and Rustenburg needed to be made to ensure that the plant would be able to survive the new conditions. These comparisons showed that Rustenburg receives on average, 484 mm less rain per year than Barberton, indicating that irrigation was required when the Rustenburg sites were planted out with B. coddii, in order to reduce water stress. Rustenburg was also found to be, on average, 4.6°C warmer than Barberton, but as B. coddii growth responds to wet/dry seasons, as opposed to hot/cold seasons, it was not felt that this temperature difference would have a negative effect on the growth of the plants. The soil comparisons showed the contaminated Rustenburg sites to be serpentine-like in nature, with respect to Barberton, again giving confidence that the plant would adapt to the conditions occurring at the contaminated sites. However, to ensure optimal growth, nutrient experiments were also performed on B. coddii to ascertain the ideal macronutrient concentrations required, without inhibiting nickel uptake. These trials indicated that the individual addition of 250 mg/l ammonium nitrate, 600 mg/l calcium phosphate, 2 000 mg/l calcium chloride, 600 mg/l potassium chloride and 250 mg/l magnesium sulphate enhanced plant growth and nickel uptake, suggesting that, for phytoremediation purposes, these nutrients should be added to the medium in which the plants are growing. The growth-cycle of naturally occurring B. coddii plants in Barberton was also studied in order that seedlings could be germinated, in greenhouses, at the correct time of year so that the plants could be sown as the naturally occurring plants were germinating. From this information the seeds of the plants could be collected at the correct time of year and the above ground biomass harvested when the nickel concentrations were at their highest. It was found that the plants began to germinate as the first rains fell, which was generally at the beginning of September, and plant maturity was reached at about five months, after which flowers were produced. Seeds were produced from the flowers and these matured and were wind-dispersed one month to six weeks after full bloom, usually during February. The plants then started to die back and dry out and dormancy was reached about nine months after germination, generally in about mid- to late- May. It was found that the nickel concentration was at its highest about one month after the plants had begun to dry out and thus it was decided that the above ground biomass would usually be harvested at the end of April each season, in order to achieve maximum nickel recovery. Finally, in order that the plant's potential for use in phytoremediation could be fully assessed, field trials at the contaminated sites in Rustenburg were performed. Germination procedures were developed for the mass production of B. coddii and it was found that, although fully formed plants could be propagated in tissue culture, it was cheaper and faster to germinate the seeds in speedling trays, containing a zeolite germination mix, in greenhouses. It was also found that the seeds had a low germination rate, due to dehydration of the embryos and thus, in order to obtain the number of plants required, four to five times the amount of seeds needed to be sown. The two-month-old seedlings were transferred to potting bags, containing a mixture of potting soil and RBMR soil, and grown up in the greenhouse for a further three months. This growth period allowed B. coddii to adapt to the RBMR soil and also ensured that the plants were relatively healthy when transplanted into three prepared sites at RBMR. The plants were allowed to grow for the entire season after which the above ground biomass, comprising the leaves and stems, was harvested, dried and then ashed in an ashing vessel designed by the author, with the help of Mr K Ehlers. The ashed material was acid-leached with aqua regia in order that the base metals (mainly nickel) and precious metals could be removed from the silicates and carbonised material. The acid solution was then neutralised, causing the base metals (mainly nickel) and precious metals to be precipitated. This precipitate was then smelted with a flux in order that nickel buttons could be formed. Thus, from all the phytoremediation trials it was found that this process is highly successful in employing B. coddii for the clean-up of nickel-contaminated sites. This constitutes the first time that such a complete phytoremediation process has ever been successfully developed with B. coddii as the phytoremediation tool. It also appears to be the first time that phytoremediation has been performed "commercially" to produce a saleable metal product. The success of this project has stimulated Amplats to continue with, and expand it, to include more studies on phytoremediation as well as in the biomining of certain areas containing very low levels of precious metals which, with conventional techniques, were previously not worth mining. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1998.

A final cover on a waste impoundment is the main physical barrier between the waste impoundment and the environment designed to protect against physical, chemical and biological factors isolating the waste from the atmospheric environment. Since the early 1990‟s regulators in the United States have started accepting vegetation covers in lieu of the prescriptive covers. Currently in South Africa, data that provide field performance comparisons of alternative vegetation covers are few or non-existent; hence a research program was undertaken by an industrial corporation in South Africa to determine the potential use of vegetation covers. In proposing a practical way forward, the Company (AECI Limited) reached an understanding with the Regulators that a vegetated evapotranspiration (ET) cover, would be acceptable provided that its performance in limiting surface water infiltration (and subsequent leaching) could be quantitatively demonstrated. The overall object of this research was to determine the capability of vegetation cover to limit effluent leaching from a waste impoundment. Analysis of the following sub-objectives were required to address and give answers to this study (1) determine, as accurately as possible a climatic water balance on the vegetation covers, (2) determine the geohydrological properties of the material of the waste impoundment, (3) determine the fate of the water i.e. proportion reused via evapotranspiration as opposed to the proportion infiltrating the waste body beneath the root zone and (4) determine the leaching potential below the waste. The study identifies and evaluates the climatic (above ground) and geohydrological (sub-surface) parameters used to estimate the water balance of the materials for a waste impoundment. The study then utilizes these parameters at the respective sites in a finite-element model, called the HYDRUS-2D model, to simulate the water balance of the material. The simulated water balance results were then compared against collected field data, which provide the evidence of the efficiency of a vegetation cover to limit effluent from the impoundment. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2009.

AtPAP15 is one of the purple acid phosphatases expressed by Arabidopsis thaliana that has been extensively studied. Purified AtPAP15 has been shown to exhibit both phytase and phosphomonoesterase activities in acidic pH with maximal activity at pH 4.5. AtPAP15 is a phosphorus starvation inducible (PSI) gene that is expressed highly during phosphorus deficient conditions. In the current study, AtPAP15 was overexpressed in Nicotiana tabaccum under cauliflower mosaic virus (CaMV35S) constitutive promoter. After PCR confirmation of the gene, plants were transferred to the greenhouse and allowed to grow in pots. The pots contained Sta-Green potting mix (Lowe’s Inc., Mooresville, North Carolina,U.S.). Individual flowers were covered to ensure selfpollination. Seeds that were generated (T1 generation) were used for functional analyses. The T1 seeds were germinated in different phosphorus conditions- phosphorus deficient (0mM), optimum inorganic phosphorus (1.25mM), high inorganic phosphorus (20mM) and high organic phosphorus (20mM AMP and IHP)- and analyzed for total biomass, primary root length, soluble phosphorus (Pi) content and acid phosphomonoesterase (Apase) activity. Results demonstrated that under high organic phosphorus conditions, transgenic lines presented increased biomass, longer primary root length, high total soluble phosphorus and high Apase activity than the wild-type. These findings will have important implications for soils with high phosphorus content since most phosphorus in soil is available in organic form. As part of this thesis, Agrobacterium-mediated in planta transformation protocol for transformation of Medicago sativa (alfalfa) was standardized. The protocol would be used to develop transgenic alfalfa expressing AtPAP15. In this protocol, shoot apical position of 2d old seedling is excised followed by infection by Agrobacterium tumefaciens strain EHA 105 for TDNA transfer. The construct used was pCAMBIA1302 harboring mGFP reporter gene. The construct also contained kanamycin, rifampicin and chloramphenicol selection markers. Confirmation of mGFP expression in the transformed alfalfa seedling was performed using Zeiss microscope fitted with GFP filters. Results showed that this protocol is replicable and efficient. In planta transformation method, therefore, may be used for the development transgenic alfalfa.

Evaluation of heavy metal accumulation in soils and plants is of environmental importance due to their health effects on humans and other biota. Phytoremediation is an emerging technology that can stabilize or remove heavy metals in soil utilizing metal tolerant plant species (metallophytes). The objectives of this study were to: (1) assess a Superfund site, and a lead-acid battery dump for concentrations of soil lead (Pb) and cadmium (Cd); (2) assess the various chemical fractions of Pb and Cd at both sites; (3) identify native herbaceous vegetation surviving in the contaminated soils; and (4) evaluate plant uptake of Pb and Cd. Soil Pb and Cd concentrations at the Superfund site averaged 55,480 and 8.5 mg/kg, respectively. Soil Pb at the Superfund site occurred primarily in the carbonate, sulfide/residual, and organic chemical fractions (41.6, 28.6, and 26.7, respectively). Soil Pb and Cd concentrations at the dump site averaged 29,400 and 3.9 mg/kg, respectively. Soil Pb occurred mostly in the organic and carbonate fractions as 48.5 and 42.5% respectively. Plant uptake of Pb at both sites ranged from non-detectable (Agrostemma githago, Plantago rugeli, Alliaria officinalis shoots), to 1800 mg/kg (Agrostemma githago root). Cd uptake was maximal in Taraxacum officinale at 15.4 mg/kg (Superfund site). In the majority of plants studied, root Pb and Cd content was higher than for shoots (>_65%). Tissue and soil fractions yielded r2 of 0.61 and 0.57 for the soluble and carbonate-Pb fractions, respectively. The plants with the greatest tissue concentrations of Pb and Cd were predominantly herbaceous species, some of which produce sufficient biomass to be practical for phytoremediation technologies. / Department of Natural Resources and Environmental Management

Phytoextraction is an innovative method that can be implemented to remove lead (Pb), cadmium (Cd), and zinc (Zn) from contaminated soil. A two-year field and growth chamber study was conducted to determine the effectiveness of several soil amendments in the phytoextraction or stabilization of Pb, Cd and Zn. Field plots were established at the Memorial Drive Dump site (Muncie, IN). Amendments included composted municipal solid waste (MSW) (25 Mg/ha), dried sewage sludge (SS) (25 Mg/ha), citric acid (2 mmol/kg soil), ethylenediaminetetraacetic acid (EDTA) (2 mmol/kg soil), and a control. A mixed grass crop was capable of survival and growth in the contaminated soil; in contrast, Helianthus annuus (sunflower) and Ambrosia artemisiifolia (ragweed) failed to become established. Soil Pb and Zn occurred primarily in the carbonate, organic-bound and residual forms (23.1%, 31.8%, and 44.4%, Pb, respectively and 11.4%, 26.5%, and 60.2% Zn, respectively). Soil Cd occurred primarily in the soluble, carbonate, and residual forms (21.8%, 21.3%, and 27.4%, respectively). In a growth chamber leaching study, soil Pb was most rapidly leached from the 0.1 M EDTA treatment (51.7 mg Pb over 200 pore volumes); the 0.001M resulted in the lowest leaching. Soil Pb was most rapidly leached using 0.1 M HEDTA (52.1 mg Pb over 200 pore volumes). The 0.001 M HEDTA rate resulted in lowest Pb leaching. Addition of an NPK solution resulted in minimal Pb leached. The MSW and dried sludge treatments resulted in the greatest dry matter production on the plots. Dry matter production was significantly (p >0.05) higher in the MSW (1.09 MT/ha) and SS (95 MT/ha) treatments. Addition of EDTA, citric acid and MSW significantly (p < 0.05) increased the amount of Cd uptake into plant tissue. The study also demonstrated the ability of vegetation to establish and grow on toxic soil and to uptake Pb, Cd, and Zn. / Department of Natural Resources and Environmental Management

Plant species were assessed for recolonization of a brownfield in Muncie, IN. In a greenhouse study, soil was seeded to perennial ryegrass (Lolium perenne), red clover (Trifolium pratense) and sunflower (Helianthus annuus). Selected pots were amended with leaf compost and mycorrhizal fungi. Soil and plant tissue were analyzed after 30 and 90 days. Ryegrass and compost were studied at the brownfield site. In the greenhouse, red clover was capable of concentrating the greatest quantity of Cd, Cu, Cr, Ni, and Pb in above-ground biomass (all soil treatments combined). Compost + mycorrhizal fungi treatment resulted in highest Cd, Cu, and Zn plant concentrations (all plant treatments combined). Compost resulted in the highest tissue Cr and Ni concentrations. The reported study demonstrates that this brownfield is capable of being revegetated by turf and legume species. Each infertile and/or toxic site must be assessed for revegetation species on a case-by-case basis. / Department of Natural Resources and Environmental Management

As the third most important greenhouse gas, nitrous oxide (N2O) is a stable greenhouse gas and also plays a significant role in stratospheric ozone destruction. The primary anthropogenic source of N2O stems from the use of nitrogen in agriculture, with soils being the major contributors. Currently, the annual N2O emissions from this “soil–microbe-plant” system is more than 2.6 Tg (one Tg equals a million metric tons) of N2O-N globally. My doctoral studies aimed to explore innovative strategies for N2O mitigation, in the context of environmental microbiology’s potential contribution to alleviating global warming. The bacterial enzyme nitrous oxide reductase (N2OR), naturally found in some soils, is the only known enzyme capable of catalyzing the final step of the denitrification pathway, conversion of N2O to N2. Therefore, to “scrub” or reduce N2O emissions, bacterial N2OR was heterologously expressed inside the leaves and roots of transgenic plants. Others had previously shown that the functional assembly of the catalytic centres (CuZ) of N2OR is lacking when only nosZ is expressed in other bacterial hosts. There, coexpression of nosZ with nosD, nosF and nosY was found to be necessary for production of the catalytically active holoenzyme. I have generated transgenic tobacco plants expressing the nosZ gene, as well as tobacco plants in which the other four nos genes were coexpressed. More than 100 transgenic tobacco lines, expressing nosZ and nosFLZDY under the control of rolD promoter and d35S promoter, have been analyzed by PCR, RT-PCR and Western blot. The activity of N2OR expressed in transgenic plants, analyzed with the methyl viologen-linked enzyme assay, showed detectable N2O reducing activity. The N2O-reducing patterns observed were similar to that of the positive control purified bacterial N2OR. The data indicated that expressing bacterial N2OR heterologously in plants, without the expression of the accessory Nos proteins, could convert N2O into inert N2. This suggests that atmospheric phytoremediation of N2O by plants harbouring N2OR could be invaluable in efforts to reduce emissions from crop production fields.

Thesis (Ph.D.) -- University of Western Sydney, Macarthur, 2001. / A thesis presented to the University of Western Sydney, in partial fulfillment of the requirements for the degree of Doctor of Philosophy, December, 2001. Bibliography : leaves 163-173.

Thèse (D.R.Min.) -- Université du Québec à Chicoutimi, programme en association avec l'Université du Québec à Montréal, 2006. / Bibliogr.: f. 150. Document électronique également accessible en format PDF. CaQCU

Submitted by Maria Medeiros (maria.dilva1@ufcg.edu.br) on 2018-08-30T13:13:44Z No. of bitstreams: 1 ELIS GEAN ROCHA - DISSERTAÇÃO (PPGECA) 2018.pdf: 5120765 bytes, checksum: 11074ea88b01e31806a1d0befc8974ef (MD5) / Made available in DSpace on 2018-08-30T13:13:44Z (GMT). No. of bitstreams: 1 ELIS GEAN ROCHA - DISSERTAÇÃO (PPGECA) 2018.pdf: 5120765 bytes, checksum: 11074ea88b01e31806a1d0befc8974ef (MD5) Previous issue date: 2018-02 / Capes / As águas urbanas passam por diversas intervenções no processo de desenvolvimento das cidades, sofrendo, muitas vezes, impactos negativos devido à ausência de planejamento e saneamento básico. A eutrofização, causada pelo enriquecimento dos corpos hídricos por nutrientes advindos de despejos de esgotos sem tratamento é um dos grandes problemas das águas superficiais urbanas. Tendo em vista a necessidade do desenvolvimento sustentável das cidades, continuamente procura-se por alternativas para o tratamento de águas poluídas que possam atuar de forma harmônica com a paisagem urbana. Este trabalho teve como objetivo a construção de um sistema de jardins flutuantes para a remediação das águas do pequeno açude localizado no campus sede da Universidade Federal de Campina Grande – PB. O experimento foi realizado numa área experimental de 400 m² de espelho d’água, a montagem do sistema se dividiu em três etapas, atingindo no final da terceira etapa uma cobertura superficial de 10%. As estruturas modulares foram construídas com bambu e paletes, sendo utilizadas diferentes espécies de plantas aquáticas e ornamentais e substrato de fibra de coco. O monitoramento foi feito por coletas semanais, em 13 pontos distribuídos na área experimental. Os parâmetros analisados foram: temperatura, pH, turbidez, condutividade elétrica, OD, DBO, DQO, clorofila-a, fósforo total, ortofosfato solúvel. Em relação às estruturas dos jardins, aquelas que não apresentaram problemas de estabilidade e flutuação, tiveram bons resultados e proporcionaram o bom desenvolvimento das plantas. As análises estatísticas dos dados mostraram que na área em estudo, houve variações significativas em todos os parâmetros estudados, apresentando resultados positivos de remoção de matéria orgânica e nutrientes em alguns dos dias monitorados. As interferências ambientais na área em estudo, como a precipitação e as cargas de efluentes, dificultaram a análise de eficiência dos jardins na qualidade da água, mas, apesar disso, foram encontrados resultados importantes. / Urban waters go trough many intervencions during the process of development of cities. Many times suffering negative impacts due to absence of planning and basic sanitation. Eutrophication caused by enrichment of water bodies by nutrientes from sewage dumping without treatment is one of the great problems of superficial urban waters. Because of the need for sustainable development in cities and the search for alternatives for the treament of polluted waters, this study has the objective of constructing a system of floating gardens to treat the waters of the pond located on the main campus of the Federal University of Campina Grande, Paraiba, Brazil. The experiment was performed on área, area of 400m2 of the superficial lawyer of the the pond. The building of the system was divided into three stages; at the end of the third stage, 10% of the superficial area was covered, modular structures were built with bamboo and pallets. Different species of ornamental aquatic plants and cocunut fiber substrate were used. Monitoring was performed weekly by collections from 14 points distributed throughout the experimental área. The parameters analyzed were; temperature, pH, turbidity, electric conductivity, DO, BOD, COD, clorophyl-a, total phophorus, soluble orthoposphate, TNK, amoniacal nitrogen, nitrite, nitrate, TDS, VDS, FDS. The structures of the gardeens that did not present problems with stability and flotation had good results and promoted good development. Statistical analysis of the data revealed in the study area, there were significant variations in all the studied parameters, presenting positive results of organic matter and nutrients removal in some of the monitored days. As environmental interferences in the study area, such as precipitation as effluent loads, made it difficult to analyze the efficiency of gardens in water quality, but despite this, important results were found.