3. Electronic Theses and Dissertations (ETDs) - All submissions
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Item The rhizobiome of Vachellia (Acacia) woodlands surrounding Witwatersrand gold and uranium mine tailings(2018) Buck, Michelle ToniPhytoremediation of mine tailings and surrounding areas provide the most cost-effective means of alleviating their pollutant effects. Research has shown that successful revegetation of mine tailings and surrounding areas can be optimised by providing appropriate microbial symbionts for the plants. Microorganisms are beneficial to plant growth and health which is essential for revegetation and phytoremediation of contaminated sites. The aim of this study was to assess the microbial status of Vachellia karroo rhizosphere currently growing on the surrounding areas of two mining operations, namely Vaal River (VR) and West Wits (WW) Operations. Metataxonomy is the study/use of genetic material isolated from field/environmental samples. It has been reported from metataxonomy studies that a large amount of microbial biodiversity had been missed by cultivation-based methods. Metataxonony allows for an unbiased genetic analysis from all members of the sampled community since it is a PCR directed sequencing approach for identification. Root and soil samples were collected in spring, roots were removed and physio-chemical analyses were preformed on the soil, including pH, conductivity, cation exchange capacity and XRF analysis. The soil samples were then prepared for DNA extraction by mixing/bulking 5 g of soil for each niche site and control. Then 500 mg of soil was used to extract DNA using the DNeasy® PowerSoil® Kit (Qiagen). Target genes, namely the 16S rRNA V1-V3 hypervariable region and the ITS2 hypervariable region, were sequenced at Molecular Research LP MR DNA (http://www.mrdnalab.com), Texas. The rhizosphere bacterial communities of V. karroo from VR Operations was predominantly comprised of Acidobacteria, Actinobacteria, Bacteroidetes, Verrucomicrobia, Firmicutes, Planctomycetes, Gemmatimonadetes, (between 1 % and 47 % depending on the niche sites and controls) and the remainder of the phyla (Chloroflexi, Armatimonadetes, Nitrospirae, Candidatus Saccharibacteria, candidate division WPS-1 and candidate division WPS-2) were less than 1 %. The numbers all represent the average percentage of sequences across 14 phyla. For WW Operations the bacterial communities were predominantly comprised of Actinobacteria, Acidobacteria, Planctomycetes, Verrucomicrobia, Proteobacteria, Bacteroidetes, Chloroflexi, Firmicutes, Gemmatimonadetes, (between 0.7 % and 41.4 % depending on the niche sites and controls) and the remainder of the phyla (candidate division WPS-1, candidate division WPS-2, Armatimonadetes, Candidatus Saccharibacteria, Nitrospirae and Cyanobacteria/Chloroplast) were around 1 %. The numbers all iii represent the average percentage of sequences across 15 phyla. The rhizosphere fungal communities of V. karroo from VR Operations was predominantly comprised of Ascomycota (Sordariomycetes, Dothideomycetes, Eurotiomycetes, Ascomycota_unidentified, Leotiomycetes and Incertae_sedis_14); Zygomycota (Incertae_sedis_10); Basidiomycota, Agaricomycetes, Atractiellomycetes, Tremellomycetes (between 0.01 % and 77.1 % depending on the niche sites and controls) and to a lesser degree Glomeromycota (Glomeromycetes) and Fungi_unidentified (between 0.2-6 % depending on the niche sites and controls). The numbers all represent the average percentage of sequences across 12 classes. For WW Operations the fungal communities were predominantly comprised of Ascomycota (Sordariomycetes, Dothideomycetes, Eurotiomycetes, Ascomycota_unidentified, Leotiomycetes and Archaeorhizomycetes), Zygomycota (Incertae_sedis_10), Basidiomycota, Agaricomycetes, Tremellomycetes, Atractiellomycetes, Basidiomycota_unidentified, Fungi_unidentified and Glomeromycota (Glomeromycetes) (between 0.01 % and 91.42 % depending on the niche sites and controls). The numbers all represent the average percentage of sequences across 13 classes. The study represents a first report utilising metataxonomic tools in the analysis of the rhizobiome of V. karroo from areas around mine tailings. The results will assist in making decisions about future microbial surveys and applying microbial inoculum in revegetation of mine waste sites and the affected areas.Item Status and molecular identification of arbuscular mycorrhizal (AM) fungi associated with Acacia spp. on rehabilitated gold and uranium mine tailings(2015-02-04) Buck, Michelle ToniPhytoremeditation of mine tailings provides the most cost-effective means of alleviating their pollutant effects. Research has shown that successful revegetation of mine tailings can be optimised by providing appropriate microbial symbionts for the plants. The aim of this study was to assess the arbuscular mycorrhizal (AM) status of trees currently being used for phytoremediation trials of mine tailings in the Welkom gold fields, and to determine the AM fungal diversity of these sites. The Acacia spp. analysed were growing on rehabilitated gold and uranium mine tailings which had undergone different rehabilitation regimes. Planted acacia trees which had been inoculated with crude AM fungal inocula were present on one mine tailing site as compared to the second mine tailing site on which the acacias were naturally colonisers and the site had been ameliorated with garden refuse. Root and slime samples were collected in early spring and half if each initial sample was used immediately for colonisation analysis and to identify AM fungi through molecular analysis of the small subunit rRNS gene sequences; the other half of each sample was used to produce trap cultures which were used later for colonisation and molecular analysis. Total AM fungal colonisation of initial samples for planted acacies was 19 % and for naturally colonising acacias was 66 %. The total AM fungal colonisation of trap culture samples for planted acacias increased to 32 % and for naturallhy colonising acacias it increased to 78 %. Spore counts of initial samples averaged 402 spores per 100 g-1 soil for planted acacias and 455 spores per 100 g-1 soil for naturally colonising acacias. For trap culture samples, spore counts decreased by approximately 50 %. The AM fungi identified fell within 8 genera, namely, Diversispora, Rhizophagus, Scutellospora, Claroideoglomus, Cetraspora, Sclerocystis, Glomus and Redecker. The study represents a first report utilising molecular biosystematics with AM fungal DNA from colonised roots as the template. The results will assist in making decisions about future AM fungal surveys and applying AM fungal inoculum in phytoremediation trials of mine waste sites. Key words: Phytoremediation, mine tailings, arbuscular mycorrhizal (AM) fungus, Acacia, molecular identification, SSU rRNA gene sequence