Clean laboratory technology and pure culture techniques were combined to determine the effect of ectomycorrhizae on the uptake of lead by Pinus sylvestris L. seedlings. By culturing the ectomycorrhizal fungus, Pisolithus tinctorius (Pers.) Coker and Couch, in liquid Hagem's medium with different concentrations of lead (0, 50, 250 and 500 ng/g sol), it was determined that the fungus readily takes up lead from such and aqueous medium. It was also determined that the pH of the medium controls the amount of lead in solution that can be taken up by the fungus. A series of growth studies where Pisolithus was cultured on Hagem's agar indicated that the growth of the fungus is reduced when the lead concentration in the medium is approximately 25 ug/g sol. It was estimated that a lead concentration of approximately 1,000 ug/g sol may completely inhibit the growth of the fungus. Furthermore, it was determined that the acetate ion is inhibitory to the growth of Pisolithus. By culturing mycorrhizal and non-mycorrhizal seedlings of P. sylvestris and analyzing the seedlings' roots, stems and leaves for their lead concentrations, it was determined that ectomycorrhizae facilitate the uptake of lead. Ectomycorrhizal roots can take up more lead from a vermiculite/hydroponic solution mixture because of their greater surface area and exploration of vermiculite surfaces as well as their increased production of lead solubilizing acids. This additional lead found in mycorrhizal roots, however, does not appear to be translocated to the above ground portions of the seedlings. This increased uptake of lead by mycorrhizal roots is expected to take place in natural soil systems as well. / Master of Science

The ectomycorrhizal communities of three aspen-covered sites in southwestern Montana and southeastern Idaho with varying stand age, conditions, and soil types were compared. In all, 39 species of ectomycorrhizal fungi were associated with P. tremuloides. Dominant fungal species varied by site. Species reported on acidic soils in Europe and other species reported to be "early colonizers" were found exclusively on the acidic soils of the Butte site. Many "late stage" fungi such as Amanitas and Cortinarii were found on the older and more productive sites. Nine isolates of ectomycorrhizal fungi were tested for their ability to form mycorrhizae with P. tremuloides. Amanita muscaria v. formosa, A. pantherina, Paxillus vernalis, and Pisolithus tinctorius formed mantles and Hartig nets. Inocybe lacera and Piloderma croceum formed mantles but no Hartig nets. Dry weight, stem diameter, height and number of roottips increased in the young aspen seedlings inoculated with a majority of the fungi listed above. The implications for aspen survival and growth are discussed. / Master of Science

This study examined the rehabilitation potential of AM fungi with organic and inorganic fertilisers under pot and field trial conditions as well as their interaction with rhizospheric organisms and specific functional groups. In addition, the study highlighted the effects of land-use management on AM fungal populations in soil and the mycorrhizal status of some selected plants from one of the study sites. The study focussed on two sites that differ in operational activities and these included a mined area that was to be rehabilitated and a commercial farming site. A pot trial was conducted using an overburdened soil resulting from kaolin clay mining. Pots were seeded with Cynodon dactylon and treated with either Organic Tea or NPK (3:1:5) fertiliser, with or without AM fungal inoculum. The compatibility of these fertilisers with AM fungi was assessed by plant growth and percentage root colonisation. Maximum shoot height and plant biomass were observed at the 28th week with NPK (3:1:5) fertiliser supporting mycorrhizal colonisation by 80%. The result indicated the potential of AM fungi to be used in rehabilitation with minimal phosphate fertiliser. Similarly, a field trial was set-up using 17 x 17 m[superscript 2] plots in the mining site that were treated with the same organic and inorganic fertilisers as well as with AM fungal inoculum in different combinations. The interaction between AM fungi and soil microbial population was determined using culture dependent and culture independent techniques. The culture dependent technique involved the use of soil dilution and plating on general purpose and selective media. The result showed that there was no change in the total culturable bacterial number in the untreated and AM fungal treated plots, while a change in species composition was observed in the functional groups. Different functional groups identified included nitrogen fixing bacteria, pseudomonads, actinomycetes, phosphate solubilisers and the fungal counterparts. Gram-positive bacteria were observed as the predominant phenotypic type, while nitrogen fixers and actinomycetes were the predominant functional groups. Species identified from each functional group were Pseudomonas fulva, Bacillus megaterium, Streptomyces and actinomycetales bacteria. Meanwhile, fungi such as Ampelomyces, Fusarium, Penicillium, Aspergillus, Cephalosporium and Exserohilium were identified morphologically and molecularly. Furthermore, the mining site had a significantly higher bacterial number than the farming site thereby indicating the effects of land-use management on culturable bacterial numbers. The culture independent technique was carried out by cloning of the bacterial 16S rDNA and sequencing. Identified clones were Bradyrhizobium, Propionibacterium and Sporichthya. A cladogram constructed with the nucleotides sequences of identified functional species, clones and closely related nucleotide sequences from the Genbank indicated that nucleotide sequences differed in terms of the method used. The activity and establishment of the introduced AM fungal population was determined by spore enumeration, infectivity assay, percentage root colonisation and assessment of glomalin concentrations. The results indicated that the two land use types affected AM fungal populations. However, the establishment of AM fungi in the farming site was more successful than in the mining site as indicated by the higher infectivity pontential. Selected host plants, which were collected around the mine area, were observed to be mainly colonised by AM fungi and these were identified as Pentzia incana, Elytropappus rhinocerotis, Euphorbia meloformis, Selago corymbosa, Albuca canadensis and Helichrysum rosum. These plant species were able to thrive under harsh environmental conditions, thereby indicating their potential use as rehabilitation host plants. Generally, the findings of this study has provided an insight into the interaction between arbuscular mycorrhizal fungi and other soil microorganisms in two fields with differing land use management practices.

The symbiotic unit of an arbuscular mycorrhizal fungus and its host is able toachieve and maintain far higher inflow of nutrients than non-mycorrhizal roots. The colonization strategy of the mycobiont within the plant is intrinsic to the symbiosis with respect to both structural adaptations and nutrient exchange. An investigation into the effect of Glomus etunicatum colonization on the structure and phloem transport in Eragrostis curvula (Umgeni) allowed for greater insight into the dynamic of the symbiosis. The combined use of stains (such as Trypan Blue, Chlorazol Black, Safranin and Fast Green), and techniques, (such as freeze-microtome transverse sectioning and permanent slide preparations) contributed to a successful general observation of an intermediate colonization strategy using light microscopy methods. However, clarity into structural detail of mycorrhizal forms required electron microscopy studies. The SEM method used with freeze fracture was a relatively quick and simple method allowing for the observation of surface and internal features. The TEM method allowed for highresolution images providing insight into the variations in the apoplasmic compartmental form, and how this may relate to the function of the symbiosis with regard to fungal coils or arbuscules. The apoplasmic nature of mycorrhizas was substantiated and no symplasmic connections were found between symbionts. Fluorescence studies demonstrated that 5,6-carboxyfluorescein was transported through the phloem into the roots of E. curvula, but remained predominantly in the root phloem. Unloading only occurred in optimal nutrient exchange areas of meristimatic lateral or apical growth regions. It was not possible, using fluorescence techniques and related equipment available, to conclusively establish if there were symplasmic connections between the mycobiont and its host or if bidirectional transfer of nutrients occurred at the same interface.

Arbuscular mycorrhizal (AM) fungi form an obligate symbiosis with many wild and cultivated plants. The plant may benefit from improved nutrient uptake (particularly phosphorus) and resistance to drought, pests and disease. Compared to more natural habitats, arable systems support reduced diversity of AM fungi, with community structure shifted in favour of ruderal types. Physical disturbance is an important driver of these differences and reduced tillage systems might facilitate greater utilisation of the AM symbiosis as part of more sustainable production systems. In this study, the structure of root-associated AM fungal communities in barley grown under a range of tillage regimes was characterised, using high-throughput molecular methodology. AM fungal community structure was significantly influenced by tillage intensity, with soils subject to high tillage burden richest in ruderal types. These findings were consistent in both winter and spring barley cultivars. Significant temporal changes in AM fungal community structure suggested an important role for plant growth stage in determining AM fungal community dynamics. Functional differences can occur between fungal species and different fungus-plant combinations, and in the second part of this study the functional consequences of changes in community structure due to tillage were tested in a model system using intact field-soil cores. There were clear differences in AM fungal community structure due to differential physical disturbance. Although no functional effects were observed, this may have been due to the inherent limitations of recreating field conditions in microcosm experiments. The results of this comprehensive assessment of fine-scale spatial variation in AM fungal community structure in an arable system identify tillage as an important driver of AM fungal community dynamics, and plant growth stage is also a key factor which has rarely been addressed. Elucidating the functional significance of these changes remains essential in order to justify future changes to arable management practices.

A Thesis, submitted to the Faculty of Science, University of the Witwatersrand, in fulfilment of the requirements for the degree of Doctor of Philosophy. Johannesburg, 2013 / A novel holistic approach was used to study the mycotrophic nature of commercial sugarcane varieties grown in KwaZulu-Natal, South Africa. All five varieties were mycotrophic, but N12 had the highest overall mycorrhization and was selected for a pot study to assess the growth response of sugarcane to inoculation with indigenous arbuscular mycorrhizal (AM) fungi and microflora. The pot study suggested that sugarcane will respond positively to inoculation with AM fungi, but the effects are most clear in the early phase of growth and less obvious in later elongation phases. This observation, taken together with the ability of sugarcane to grow well in sterile soil without microflora additions suggests that the plant may be facultatively mycotrophic. A multivariate analysis determined the nutrient relationships between soil and corresponding leaf nutrient levels on 72 sugarcane field plants, categorised according to either high or low percentage colonisation. Highly colonised plants were found to have more positive nutrient correlations compared to lower percentage colonised plants. AM fungi were identified from spore morphology and associated mycorrhizal bacteria (AMB) were identified by 16s rDNA analysis. Partial molecular identification was conducted using a universal eukaryotic forward NS31 primer and general fungal AM1 primers confirming the spores to be of AM fungi origin. A nested PCR was performed, using the universal fungal primers, NS5 and ITS4, followed by primer combinations to target sequences of specific Glomalean groups. Only partial molecular identification was conducted, as RFLPs were not successfully optimised. DNA from the Acaulospora gerdemannii/Acaulospora trappei group, Glomus occultum/Glomus brasilianum group, Glomus mosseae/Glomus intraradices group, Glomus etunicatum/ Glomus clariodeum group and Acaulosporaceae sensu stricto were detected, indicating AM fungi diversity. Bacteria, Brevibacillus reuszeri isolated from Scutellospora nigra, Bacillus megaterium and Stenotrophomonas maltophilia isolated from Glomus geosporum, Paenibacillus chitinolyticus and Bacillus cereus isolated from Acaulospora mellea and Gigaspora margarita spores respectively, were tested for biocontrol capability against pathogenic nematodes of Paratrichodorus, Meloidogyne and Pratylenchus genera. Meloidogyne was the least susceptible to associated mycorrhizal bacteria biocontrol and Paratrichodorus the most susceptible. These studies have contributed to understanding the role of AM in sugarcane agriculture in South Africa. Keywords: arbuscular mycorrhiza, sugarcane varieties, multivariate analysis, nutrient relationships, associated mycorrhizal bacteria, biocontrol, nematodes

Identifying the mechanisms that prevent competitive exclusion in tropical forests is a key goal of tropical ecology. Because trees are long-lived organisms, it is complicated to test theory related to coexistence. However, the seedling stage, during which tree mortality is the highest, offers an ideal proxy to evaluate mechanisms that promote or hinder tree species coexistence. This dissertation utilizes both theory and empirical approaches to investigate two mechanisms thought to influence seedling dynamics and tree species coexistence: negative feedbacks from tree natural enemies and positive feedbacks from arbuscular mycorrhizal (AM) fungi. Tree natural enemies might promote tropical tree species coexistence by acting as agents of negative density-dependent mortality. Simultaneously, tropical seedlings associate with arbuscular mycorrhizal (AM) fungi, which can increase seedling growth and survival through enhanced nutrient and water uptake. However, little is known about the effects of AM fungi on tropical tree community dynamics. In chapter 1, I developed a theoretical model that combines competition between trees, predation by tree natural enemies, and mutualism with AM fungi, and showed that a subtle balance between negative and positive feedbacks is required to reach tree species coexistence. In chapters 2 and 3, I used empirical data collected from El Yunque (Puerto Rico) to gain a better understanding of the distribution of soil fungi and tree natural enemies in a secondary tropical forest and to test some of the assumptions of my theoretical model. In chapter 2, I found evidence that soil characteristics and the tree community were important to structure soil fungal communities, and I demonstrated long-lasting effects of past human land use. If AM fungi are important to promote tropical tree species coexistence as suggested by my theoretical model, past land use could influence tree species coexistence by altering AM fungal communities, emphasizing the need for additional studies about land use legacy effects on AM fungal communities. In chapter 3, I showed that seedlings at intermediate conspecific density and from intermediate abundance tree species, hosted the richest community of natural enemies, suggesting that negative density-dependent processes might be non-linear, and partially supporting my theoretical model. Finally in chapter 4, I investigated seedling mortality and showed that natural enemies increase seedling mortality, whereas AM fungal diversity decreases seedling mortality, counteracting the local effects of natural enemies. I also found evidence that AM fungal diversity rescues rare tree species, and natural enemies reduce survivorship of more abundant species, thereby preventing competitive exclusion. Therefore, at the community scale, AM fungal diversity and natural enemies act in the same direction, promoting tropical tree species coexistence, which is consistent with the findings of my theoretical model. In conclusion, this dissertation jointly investigated the effects of negative and positive feedbacks on tropical tree species coexistence, and demonstrated the importance of combining demographic processes that are known to occur simultaneously.

No description available.

Includes bibliographical references. / xviii, 171 leaves : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, 1994

Copies of author's previously published articles inserted. / Bibliography: leaves 184-197. / xxvi, 197 leaves, [5] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Soil Science, 1995