Root exudates include important chelating compounds and can change the rhizosphere pH by several units. These changes are essential for nutrient uptake and can also alter solubility of soil contaminants and increase plant uptake. Mild root-zone stress may increase exudation and more severe stress can damage membranes and increase root turnover, all of which increase root-zone carbon. Increased carbon from this rhizodeposition can increase microbial activity, which might help degrade contaminants. We studied the effect of three types of stress on root exudation of crested wheatgrass (Agropyron cristatum): low K+, drought, and flooding. These stresses were compared to two types of controls: 100% NO3- and high NH4+:NO3- ratio. We developed an improved axenic system to keep plants microbe-free for 70 days while analyzing exudates for total organic carbon (TOC) and organic acids. Axenic conditions were confirmed by plate counts of the leachate and microscopic observations of the leachate and roots. Optimal conditions for plant growth were maintained by monitoring temperature, light, humidity, water, O2, CO2, nutrient availability, and root-zone pH. Plants were grown in Ottawa sand that was layered by size to optimize water availability. Total organic carbon released over the 70-day growth period in mg per gram dry plant was 2.6 in the control, 2.3 in the NH4+ treatment, 3.7 in the flood and K+ stress treatments, and 4.4 in the drought treatment, which was the only treatment significantly higher than controls (p = 0.05). TOC and organic acid levels in the exudates peaked before the end of the study. The peak TOC levels, expressed as mg TOC per gram new dry plant mass, were 1.9 in the control, 3.0 in the NH4+ treatment, 2.9 in the flood, and 5.8 in the drought and K+ stress treatments. Organic acids were measured by gas chromatography-mass spectrometry (GC-MS). Malic acid was the predominant organic acid, and accounted for the majority of the TOC in the drought treatment. Oxalic, succinic, fumaric, and malonic acids accounted for less than 10% of the TOC. These data indicate that stress may enhance phytoremediation by changing root-zone exudate composition.

The rhizosphere is characterized by the presence of high microbial activities which are influenced by plant root exudates. This study examined bacterial diversity and physiological functions plants rhizosphere using both culture-dependent and culture-independent techniques of seven cultivated. Physico-chemical properties of soil samples revealed that the rhizobacteria adapted well to pH ranging from 7.5 to 9.1. Macronutrients (carbon, nitrogen, calcium, magnesium, phosphorous, potassium, sodium and iron) had a wide range of concentration between 0 to 4380.1 mg/kg. Concentrations of metal elements (cadmium, cobalt, chromium, copper and zinc) from all rhizosphere samples were below the amount of 3.1 mg/kg, indicating that the samples were free from metal contaminations. Sole carbon substrates utilization of bacteria in rhizosphere samples were measured as Average Well Colour Development (A WCD) and Group-wise Average Well Colour Development (AWCDg) patterns. At seventy two hours, there was no significant difference in AWCD patterns between bacteria in all samples and there was a significant difference in AWCDg patterns. Biochemical tests showed majority of isolates had similar physiological properties to members of Bacillus genus. All the bacterial isolates exhibited positive antifungal trait, fifteen solubilized phosphate and three had cyanide production traits during in vitro plant growth promotion assays. In vitro plant growth revealed that bacterial isolate RL1 (Bacillus licheniformis) produced the highest concentration of indole acetic acid (IAA) at 25 mg/ml. Bacterial isolate RG3 (Bacillus pumilus) had the highest amino cyclopropane carboxylase (ACC) deaminase activity indicated by the high production of α-ketobutyrate produced at 4.8 mg/ml. There were significant differences in shoot length at P ≤ 5% level of significance and there was no significant difference in the number of leaves across all three inoculated plants at P ≥ 5% level of significance. Sequence and phylogenetic analysis of identified culture-dependent bacteria revealed a homologous similarity of 94 to 100% between isolates sequences and GenBank sequences. From this, 81% of the sequences were closely related to Firmicutes, 13% to Actinobacteria and 6% to Proteobacteria. From cultureindependent method, only 8 PCR-DGGE bands were detected, the 200 bp sequences in the 16S rRNA fragment showed 91 to 100% homologous similarity to GenBank sequences. Their 16S rRNA sequences was closely related to 50% uncultured bacterium clones, 25% Firmicutes, 13% Proteobacteria and 12% Bacteroidetes sequences. Both culture-dependent and cultureindependent techniques were precise in the identification and description of bacterial community in rhizosphere. / Thesis (M.Sc) North-West University, Mafikeng Campus, 2014

No description available.

Cheatgrass is an invasive weed that has come to dominate large areas of the western United States. Once an ecosystem has been converted to a cheatgrass monoculture, it is extremely difficult to restore native vegetation. Cheatgrass negatively impacts wildlife and increases wildfire frequency and intensity. Understanding how cheatgrass so effectively invades western ecosystems is essential to turning the tide of invasion. One possible key to cheatgrass’ success is alteration of soil nutrient cycling. The goal of this study is to explore how nitrogen (N) may accumulate in cheatgrass soils via redistribution of N within soil N pools. To accomplish this we investigated soil N cycling in soils underneath cheatgrass and crested wheatgrass. We used a 15N isotope tracer to determine the contribution of root exudates to soil N pools. During the 1-week 15N tracer experiment, cheatgrass roots exuded more than twice as much N (0.11 mg N kg-1 soil d-1) as crested wheatgrass roots (0.05 mg N kg-1 soil d-1). We propose that exudation of high N content root exudates leads to the changes in soil N pool size and transformation rates commonly observed in soils under cheatgrass. This research uses a simple and relatively inexpensive isotope tracer to shed light on mechanisms by which invasive plants may alter soil processes. By understanding these mechanisms we may be able to develop strategies for better managing cheatgrass invasion.

Phytoremediation systems for petroleum hydrocarbons rely on a synergistic relationship between plants and their root-associated microbial communities. Plants exude organic compounds through their roots, which increase the density, diversity and activity of plant-associated microorganisms, which in turn degrade hydrocarbons. Understanding the mechanisms driving this relationship poses one of the more intriguing challenges in phytoremediation research. This study was designed to address that challenge. Plant-microbe interactions in a weathered-hydrocarbon contaminated soil were examined under controlled growth chamber, and field conditions. In both environments single-species grass treatments initially facilitated greater total petroleum hydrocarbon (TPH) degradation than <i> Medicago sativa </i> (alfalfa), mixed species, or control treatments. In growth chamber studies increased degradation was linked to increased aliphatic-hydrocarbon degrader populations within the rhizosphere. Under field conditions, specific recruitment of endophytic aliphatic-hydrocarbon degraders in response to high TPH levels may have facilitated increased degradation by the grass <i> Elymus angustus</i>(Altai wild rye, AWR). AWR stably maintained these communities during times of local drought, enabling them to act as subsequent source populations for rhizosphere communities. The broad phylogenetic diversity of AWR endophytes, compared to the <i> Pseudomonas</i>-dominated communities of other plants, contributed to the observed stability. The relative composition of exudates released by plants also impacted both degradation activity and potential. Alfalfa released higher concentrations of malonate, which hindered degradation by decreasing metabolic activity and concomitantly inhibiting catabolic plasmid transfer. In contrast, AWR exudates contained high levels of succinate, which was linked to increased catabolic gene expression and plasmid transfer. A reciprocal relationship between exudation patterns and endophytic community structure likely exists, and both parameters have a specific influence on rhizosphere degradation capacity. In this study, grasses were more successful in maintaining the specific balance of all parameters required for the transfer, preservation, and stimulation of hydrocarbon catabolic competency.

Phytoremediation systems for petroleum hydrocarbons rely on a synergistic relationship between plants and their root-associated microbial communities. Plants exude organic compounds through their roots, which increase the density, diversity and activity of plant-associated microorganisms, which in turn degrade hydrocarbons. Understanding the mechanisms driving this relationship poses one of the more intriguing challenges in phytoremediation research. This study was designed to address that challenge. Plant-microbe interactions in a weathered-hydrocarbon contaminated soil were examined under controlled growth chamber, and field conditions. In both environments single-species grass treatments initially facilitated greater total petroleum hydrocarbon (TPH) degradation than <i> Medicago sativa </i> (alfalfa), mixed species, or control treatments. In growth chamber studies increased degradation was linked to increased aliphatic-hydrocarbon degrader populations within the rhizosphere. Under field conditions, specific recruitment of endophytic aliphatic-hydrocarbon degraders in response to high TPH levels may have facilitated increased degradation by the grass <i> Elymus angustus</i>(Altai wild rye, AWR). AWR stably maintained these communities during times of local drought, enabling them to act as subsequent source populations for rhizosphere communities. The broad phylogenetic diversity of AWR endophytes, compared to the <i> Pseudomonas</i>-dominated communities of other plants, contributed to the observed stability. The relative composition of exudates released by plants also impacted both degradation activity and potential. Alfalfa released higher concentrations of malonate, which hindered degradation by decreasing metabolic activity and concomitantly inhibiting catabolic plasmid transfer. In contrast, AWR exudates contained high levels of succinate, which was linked to increased catabolic gene expression and plasmid transfer. A reciprocal relationship between exudation patterns and endophytic community structure likely exists, and both parameters have a specific influence on rhizosphere degradation capacity. In this study, grasses were more successful in maintaining the specific balance of all parameters required for the transfer, preservation, and stimulation of hydrocarbon catabolic competency.

Tall fescue (Lolium arundinaceum (Schreb.) Darbysh.) is a cool-season perennial grass used in pastures throughout the Southeastern United States. The grass can harbor a fungal endophyte (Epichloë coenophiala) thought to provide the plant with enhanced resistance to biotic and abiotic stress. However, the alkaloids produced by the common variety of the endophyte cause severe animal health issues resulting in a considerable amount of research focused on eliminating the toxic class of alkaloids while retaining the positive abiotic and biotic stress tolerance attributes of the other alkaloids. In doing so, very little attention has been paid to the direct influence the fungal-plant symbiosis has on rhizosphere processes. Therefore, my objectives were to study the influence of this relationship on plant biomass production, root exudate composition, and soil biogeochemical processes using tall fescue cultivars PDF and 97TF1 without an endophyte (E-), or infected with the common toxic endophyte (CTE+), or with two novel endophytes (AR542E+, AR584E+). I found that root exudate composition and plant biomass production were influenced by endophyte status, tall fescue cultivar, and the interaction of cultivar and endophyte. Cluster analysis showed that the interaction between endophyte and cultivar results in a unique exudate profile. These interactions had a small but perceptible impact on soil microbial community structure and function with an equally small and perceptible impact on carbon and nitrogen cycling in soils from rhizobox and field sites. These studies represent the first comprehensive analysis of root exudate chemistry from common toxic and novel endophyte infected tall fescue cultivars and can be used to help explain in part the observed changes in C and N cycling and storage in pastures throughout the Southeast U.S..

Toxic elements are present in polluted water from mines, industrial outlets, storm water etc. Wetland plants take up toxic elements and increase the pH of the medium. In this thesis was investigated how the shoots of submerged plants and roots of emergent plants affected the pH of the surrounding water in the presence of free toxic ions. The aim was to clarify the mechanisms by which these plants change the surrounding water pH in the presence of toxic ions. The influence of Elodea canadensis shoots on the pH of the surrounding water was studied in the presence of cadmium (Cd) at low initial pH (4-5). The involvement of photosynthetic activity in the pH changes was investigated in the presence and absence of Cd. The cytosolic, vacuolar and apoplasmic pH changes as well as cytosolic Cd changes in E. canadensis were monitored. The influence of Eriophorum angustifolium roots on the pH of the surrounding water was investigated in the presence of a combination of Cd, copper, lead, zinc and arsenic at low initial pH (3.5). Eriophorum angustifolium root exudates were analyzed for organic acids. Elodea canadensis shoots increased the pH of the surrounding water, an effect more pronounced with increasing Cd levels and/or increasing plant biomass and increased plant Cd uptake. The pH increase in the presence of free Cd ions was not due to photosynthesis or proton uptake across the plasmalemma or tonoplast. Cadmium was initially sequestered in the apoplasm of E. canadensis and caused its acidosis. Eriophorum angustifolium roots increased the surrounding water pH and this effect was enhanced in the presence of arsenic and metals. This pH increase was found to depend partly on the release of oxalic acid, formic acid and succinic acid by the plants. In conclusion, E. canadensis shoots and E. angustifolium roots were found to increase the low initial pH of the surrounding water. The pH modulation by these species was enhanced by low levels of free toxic ions in the surrounding water. / At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Submitted. Paper 3: Submitted. Paper 4: Manuscript.

Phytophthora cinnamomi is an important pathogen of many plant species in natural ecosystems and horticulture industries around the world. In Western Australia, a high proportion of native plant species are susceptible to P. cinnamomi attack. Acacia pulchella, a resistant legume species native to Western Australia has been considered as a potential biological control tool against P. cinnamomi. To develop effective control methods, it is important to understand the interactions between the control agent and the different life forms of the pathogen. In this thesis the interactions are investigated between P. cinnamomi and varieties of A. pulchella which occur in jarrah (Eucalyptus marginata) forest and sand plain ecosystems. The soil inoculum of P. cinnamomi was compared under the potted plants of the three common varieties of A. pulchella, var. pulchella, var. glaberrima and var. goadbyi. These were grown in infected jarrah forest soil in the glasshouse and in vitro in a sterilised soil-less mix aseptically. Acacia urophylla (a species non suppressive towards P. cinnamomi) was also included as a control. An isolate of the most commonly found clonal lineage of P. cinnamomi in the jarrah forest, A2 type 1 was selected for use in experiments after testing showed it reliably produced zoospores and chlamydospores both axenically and in non-sterile conditions, in comparison to several other isolates. The lowest survival of P. cinnamomi inoculum was found under A. pulchella var. goadbyi plants grown both in non sterile soil and in aseptic soil-less mix. All the life forms of P. cinnamomi were affected by A. pulchella (Chapters 2, 3, 4 and 5). The soil leachates from potted plants of A. pulchella var. goadbyi reduced sporangial production (Chapter 2) and caused cytoplasm collapse of chlamydospores (Chapter 3). The confirmation was obtained that soil under A. pulchella was inhibitory to sporangial stage of P. cinnamomi and new evidence was obtained on chlamydospore inactivation. Cytoplasm collapse in the chlamydospores was observed both for chlamydospores on mycelial discs on Mira cloth exposed to the soil leachate and within infected roots buried in soils under the three varieties of A. pulchella plants. The effect was strongest under the plants of A. pulchella var. goadbyi and indicated that the chlamydospores of P. cinnamomi are unlikely to act as persistent structures under A. pulchella var. goadbyi plants. In Chapter 4, bioassays were conducted with axenically produced mycelia, chlamydospores and zoospores to test the inhibitory effect of the root exudates collected from aseptically grown A. pulchella var. goadbyi plants. The zoospores of the same isolate used in the soil leachate tests were immobilised (became sluggish and encysted) within one to two minutes. When incubated for 24 h, zoospores predominantly clumped and germ tubes were observed only from the clumped ones. Chlamydospores produced by four isolates of the common A2 type 1 strain and the only one A2 type 2 strain available at the time were tested. A higher percentage of chlamydospores collapsed and a very low percentage germinated after 24 h. Chlamydospores of all the A2 type 1 isolates were inhibited by the root exudates whilst the A2 type 2 isolate remained viable. The findings showed that the suppressive effect must be due at least in part to substances exuded by the A. pulchella plants. However, it appeared that the A2 type 1 isolates were more vulnerable to this effect than the single A2 type 2 isolate. In Chapter 5, the effect of season on sporangial suppression of P. cinnamomi was shown using field soils collected from three jarrah forest soil vegetation types and a Banksia woodland on Bassendean sand, collected in winter and summer. The effect of age of A. pulchella plants was demonstrated using the soils collected from rehabilitated bauxite mine pits. In all the locations soils were collected under A. pulchella plants and 5 m away from the nearest A. pulchella. An effect of soil type was evident as whilst the soil leachates made from the three lateritic jarrah forest soil types where A. pulchella is common in the understorey were suppressive to the sporangial stage of P. cinnamomi, this effect was not evident in the Bassendean sand under A. pulchella. A. pulchella soils collected in winter were less suppressive towards sporangial production than soils collected in summer. An effect of plant age was demonstrated as soil leachates from four year-old A. pulchella stands in rehabilitated bauxite mine sites were more suppressive for sporangia than leachates from one year-old stands. Further information on the behaviour of the pathogen in soil and in potting mix with and without A. pulchella was obtained by infecting lupin radicles with an isolate of each A2 type, 1 and 2 strains of P. cinnamomi and burying them in the soil under the three varieties of A. pulchella plants. After a week, the chlamydospores were mostly collapsed and hyphae deteriorated. Oospores were observed and in significant numbers under the potted plants of A. pulchella var. glaberrima. Isolates of all three clonal lineages of P. cinnamomi found in Australian soil were tested for the ability to produce oospores. Two isolates of the A1 and A2 type 2 and three isolates of the common A2 type 1 were screened. The two isozyme types of the A2 clonal lineage isolated in Australia varied in ability to self and produce oospores in planta in several soils from the jarrah forest. The isozyme type 2 of the A2 clonal lineage of P. cinnamomi produced oospores under these experimental conditions. This stimulation was not effective for most of the tested isolates of the A2 type 1 and the A1 clonal lineage. The in planta oospores were viable but dormant and the oogonial-antheridial associations were amphigynous both in vitro and in vivo. For the first time it was established that, the stimulus for selfing and oospore formation in the A2 type 2 of P. cinnamomi is available in some jarrah forest soils, with and without A. pulchella and also in the potting mix used. This raises important questions for the management of the pathogen. Several factors were identified as potential stimuli for selfing. Among them, soil nutrient levels and essentially enhanced sulphur presence were found important. Temperature also played a key role. Oospores were produced abundantly at 21 – 25 ºC but not over 28 ºC. The biology of P. cinnamomi has been studied for several decades but some important aspects remain un-researched. This thesis pioneers research into the in planta selfing aspect of the pathogen in soil. It also improved the understanding of the interactions between P. cinnamomi and A. pulchella which to some extent supports use of A. pulchella as a biological control tool against P. cinnamomi. However, attention is drawn to the natural mechanisms of this complex pathogen to survive in planta by producing oospores, the most persistent form of its life cycle.

The aim of this thesis was to assess the effect of root exudates on the biodegradability of dissolved organic matter. The experiment was done in laboratory conditions, using the dissolved organic matter sampled in a spruce swamp forest located in Šumava National Park and an artificial mixture of root exudates prepared according to known composition of root exudates released by peatland plants. Main hypothesis was that the input of root exudates into the peatland pore water will affect decomposition of less-degradable dissolved organic matter, with the resulting effect dependent on the quantity and quality (C:N ratio) of the input.