Fusarium root rot is one of the most important diseases of pulse crops, with numerous Fusarium spp. comprising the disease complex. Fusarium solani and F. avenaceum have been reported to be major pathogens in the pea root rot complex, and all commonly grown varieties are susceptible. Greenhouse methods to evaluate peas for resistance to Fusarium root rot resulted in inconsistent disease severity across varieties. In 2015, F. avenaceum infested field plots were more heavily damaged based on emergence and yield than F. solani infested plots, and opposite trends were observed in 2016. Differences in root rot severity between years could be due to F. solani infestation causing more damage under warmer temperatures, while plots infested with F. avenaceum caused more damage under cooler temperatures. These results highlight the difficulties observed when screening for soil-borne pathogens, and the increased difficulties when a pathogen complex and changing environmental conditions are involved.

Phytophthora medicaginis is an important soil-borne oomycete pathogen of lucerne (Medicago sativa) and chickpea (Cicer arietinum) within Australia and overseas. To understand the host/pathogen interaction, a pathosystem was developed using the model legume Medicago truncatula. Using the resources developed for genetics and molecular characterisation in this model plant, the aim of this research was to understand the interaction between M. truncatula and P. medicaginis, with a view to improving resistance to this important pathogen in related legumes. To observe and characterise the interaction between M. truncatula and P. medicaginis, a pathosystem was developed by first screening a germplasm collection of 99 M. truncatula accessions. This revealed a continuous distribution in disease phenotypes with variable extremes in natural resistance to P. medicaginis culture UQ5750, isolated originally from M. sativa. P. medicaginis zoospore inoculation of 1-2 week-old seedlings in glasshouse experiments proved to be a robust and repeatable method to consistently confirm the responses observed for six key M. truncatula accessions; SA8618 and SA8623 exhibit high natural resistance to this pathogen, accession A17 is moderately resistant, A20 is moderately susceptible and accessions Borung and SA30199 are susceptible. To characterise the genetic basis of resistance to P. medicaginis, two reciprocal F2 populations from cross pollinations between A17 and Borung and SA8618 and SA30199 were produced and then phenotyped for disease symptoms. Genetic segregation patterns indicated the involvement of a gene with a major effect in both reciprocal populations. In particular, a 3:1 segregation ratio for resistance in the F2 populations from cross pollinations between A17 and Borung indicated the possibility of a single dominant gene for moderate resistance. Further phenotyping of F3 families is required to verify this. A M. truncatula linkage map was constructed using 50 F2 individuals of the A17 X Borung population and 49 F2 individuals from the Borung X A17 population. The map, covering 519.3 cM, is comprised of 84 SSR markers with an average distance between markers of 8.7 cM. These are evenly spaced over 7 linkage groups, including a super linkage group conferred by a translocation event between LG4 and LG8 of accession A17. Quantitative trait locus (QTL) analysis confirmed there was a QTL with a major effect in the A17/Borung reciprocal populations. A significant QTL was determined by quantifying two symptoms of P. medicaginis infection - proportion of dead/chlorotic leaves and root fresh weight. The trait loci for both symptoms were located on the same linkage group within the same region, supporting the putative position of the QTL and the authenticity of its involvement in resistance to P. medicaginis. This QTL was located on LG6 and accounted for 69.5% of the observed variation in proportion of dead/chlorotic leaves or 38.1% of the variation in root fresh weight within the inoculated populations. The effect of this QTL on resistance to P. medicaginis translated into 27.5% less dead/chlorotic leaves or 0.86 g more root fresh weight. Other QTLs with minor effects that are potentially involved in the interaction are located elsewhere on LG6 and LG2. However, the marker density of the linkage map and the population size need to be increased to verify this. In parallel to this, an F7 recombinant inbred line (RIL) population of chickpea (BG212 X Jimbour), developed by breeders at the New South Wales Department of Primary Industries (NSW DPI), was also assessed for the genetic basis of resistance to P. medicaginis. Variance component analysis of phenotype scores for this intraspecific RIL population indicated that 57.15% of the differences in between-family and withinfamily variance could be attributed to a genetic component. However, gene-based markers developed in M. truncatula and established simple sequence repeat (SSR) markers of chickpea were not sufficiently polymorphic in size to produce a linkage map for further QTL analysis. An interspecific cross between C. arietinum and C. echinospermum (Howzat X ILWC246) was also performed by breeders at the NSW DPI to develop RILs. In the duration of this research these interspecific RILs were bred to generation F3 and phenotyping assessment had not been performed. However, marker screening of the parents revealed 122 size polymorphic chickpea SSR markers. A sufficient linkage map could be produced for QTL analysis once field assessment of this population is performed. Initial screening of the M. truncatula gene-based markers on the parents of this interspecific cross also revealed that 50% show a sequence-identified base pair difference. A chickpea linkage map incorporating these markers could be comparatively mapped with M. truncatula. Molecular investigations of the M. truncatula/P. medicaginis pathosystem were performed to elucidate the possible underlying defence mechanisms involved in the observed resistance. To determine the function of ethylene in the resistant response, the characterisation of defence associated mutants of M. truncatula and Agrobacterium rhizogenes-mediated ‘hairy root’ transformations were employed. Comparison of response to inoculation of an ethylene insensitive mutant of M. truncatula (sickle) with the moderately resistant background genotype A17 showed that sickle was hypersensitive to P. medicaginis. This indicated that ethylene insensitivity was not the source of resistance to this pathogen and importantly that ethylene is a key defence signalling molecule in the moderate resistance of A17 to P. medicaginis. Agrobacterium-mediated ‘hairy root’ transformations of M. truncatula with 4GCC::Luc constructs, revealed that the production of ethylene and consequently ethylene response factors (ERFs) after inoculation by P. medicaginis was a general defence reaction by all accessions. The two susceptible M. truncatula accessions exhibited a much stronger and earlier response to inoculation than the highly resistant and moderately resistant accessions. This indicated that the resistant response may be directed by a transcriptional component governed by the host genotype, downstream of ethylene production. The M. truncatula/P. medicaginis ‘hairy root’ transformation assay has scope to be a powerful functional genomics tool for this pathogen interaction. Reverse transcriptase quantitative polymerase chain reaction (RTqPCR) was employed to determine the general patterns of gene expression and function underlying the response to P. medicaginis infection. Relative changes in gene expression of key enzymes in each of the salicylic acid, jasmonic acid, ethylene and isoflavonoid defence pathways and in genes encoding downstream target proteins revealed potential genes involved in the resistance to P. medicaginis. There was a distinct molecular difference in the response between the high and moderately resistant M. truncatula phenotypes to this pathogen. Moderate resistance to P. medicaginis in M. truncatula is possibly mediated by ethylene and involves the considerable induction of pathogenesis related protein 5 (PR5), which was not the same defence response that conferred the high resistance to P. medicaginis. Early and consistent expression of genes encoding key enzymes of the isoflavonoid pathway by the highly resistant accession indicated that phytoalexin response could be associated with the high resistance. Confirmation of the involvement of isoflavonoid phytoalexins in the high resistance response to P. medicaginis merits further investigation.

Thesis (M.S. - Hydrology and Water Resources Administration)--University of Arizona, 1987. / Includes bibliographical references (leaves 166-176).

Submitted in fulfillment of the requirements for the degree of Doctor of Technology: Health Sciences, Durban University of Technology, Durban, South Africa, 2018. / Soil-transmitted helminths are a major health concern, especially in tropical and sub-tropical regions. Poor sanitation and poverty are major pre-disposing factors contributing to increase in infections. Infection with STH is mainly through exposure to water, soil and food contaminated with the eggs of these parasites. Accurate detection and quantification of STH eggs in environmental samples is therefore critical for the determination of infection risks from exposure. Accurate detection of these eggs is also important in the adoption of risk reduction strategies. This thesis presents the development of a revised method for the accurate detection and quantification of STH eggs in different environmental matrices, such as wastewater, sludge etc. It further presents the application of this method in the comparative determination of STH egg reduction efficiencies of centralized wastewater treatment plants and decentralized wastewater treatment (DEWATS) plants in Durban, South Africa and Maseru, Lesotho. The concentration of viable STH eggs in dried sludge from Durban, South Africa and Dakar, Senegal was also determined and compared with both WHO guidelines and South African national standards for sludge reuse. The risks of infection with STHs for different populations exposed (directly and indirectly) to wastewater, wastewater contaminated surface water and sludge were determined using both quantitative microbial risks assessment and epidemiological approaches. Despite the plethora of methods available for the detection and quantification of STH eggs in the environment there is no internationally accepted method, however the most commonly used methods are based on the principles of sedimentation, differential flotation and microscopy. These are mainly adaptations of the WHO and USEPA methods. These methods were found to be similar with a few differences which affected the recovery rates reported. However, the major challenges with the conventional methods are the time needed for sample analysis and the use of reagents that could possibly affect the recovery of viable STH eggs. A new revised method was developed based on review of literature and laboratory experiments. In this method the heterogeneity of environmental samples was accounted for by the development of different pre-processing steps, involving the use of detergents to aid in the separation of eggs from particles in samples such as sludge, UD waste and untreated wastewater. Additionally, the use of sieves of different pore sizes ensured that the number of debris on the microscope slides was reduced considerably. The use of these sieves also reduced the time need for sample analysis, due to the elimination of the spontaneous sedimentation step, which is commonly used. This spontaneous sedimentation step takes between 12-24 hours therefore prolonging the time needed for sample analysis. Reagents such as acetoacetic acid and ethyl acetate were found to result in considerable loss of egg viability after just 5 minutes of exposure. This new method therefore does not involve their usage. The elimination of the use of acetoacetic acid and ethyl acetate step also reduces the number of steps involved in sample analysis. This reduces room for error as well as helping in fast analysis of samples. In addition to a much faster sample analysis the method has recovery percentages of 80.25% to 97.63% in sludge and wastewater samples respectively, with sensitivity of 2-3 eggs per liter in wastewater samples and 5-7 eggs per 20 gram of sludge. Exposure to STH eggs in the environment is mainly through wastewater, either treated or untreated, this exposure could therefore be eliminated through wastewater treatment. Centralized wastewater treatment systems are the most favored treatment options globally. These centralized treatment systems incur high cost of construction, maintenance and operations which may hamper the robustness in developing countries and rural areas. One of the most widely used alternative means of wastewater treatment is the anaerobic baffled reactors (ABRs) and planted gravel filters (PGFs) (collectively referred to as DEWATS in this thesis), which have been considered as low cost, effective wastewater treatment options. However, there is lack of comparative assessment of the STH egg removal efficiency of these two different wastewater treatment approaches. Eggs of Ascaris spp, hookworm, Trichuris spp, Taenia spp and Toxocara spp were the commonly recorded STH eggs in the untreated wastewater at the inlets of the centralized wastewater treatment plants as well as the DEWATS plants (except for Toxocara spp). There was variation in STH egg concentrations between and within the study areas, indicating difference in STH infections among the populations both in Durban and Maseru. STH egg removal varied between and within the different wastewater treatment plants as well. The DEWATS plants achieved 95-100% STH egg removals as compared to the 67 to 100% in the centralized wastewater treatment plants. This could be attributed to the difference in treatment processes. Among the different STHs, reduction in Ascaris spp eggs was significantly higher, irrespective of the type of treatment, which is attributed to the high relative density of the egg resulting in a higher settling velocity than the other STH eggs. Reduction or elimination of STH eggs through wastewater treatment is achieved by removing the eggs from the wastewater into the sludge. STH egg concentration in sludge is therefore mostly higher than in the wastewater. Sludge from Durban and Dakar after 60 days of drying under ambient environmental conditions contained very high concentration of viable STH eggs. Ascaris spp, hookworm, Trichuris spp, Taenia spp and Toxocara spp were the commonly recorded STH eggs, except for Dakar were Taenia spp and Toxocara spp were not detected in the sludge. STH egg concentrations were higher in Dakar than in Durban, with viable STH egg concentrations exceeding both the USEPA regulatory value (≤0.25 eggs/g TS) and the WHO guideline value (≤1 eggs/g TS). This variation in egg concentration could be attributed to the difference in prevalence and intensity of STH infections in the two study areas. Over a ten-month study period concentration of viable eggs in the sludge from Durban varied considerably, probably influenced by the environmental conditions. A decay rate of 0.0056 per day was calculated for egg die-off during drying. The rate of decay is low therefore drying alone cannot produce sludge meeting both local and international standards and guidelines for sludge reuse. Determination of STH infection risks due to exposure to wastewater and sludge either directly or indirectly is critical in the prevention of infection. Exposure to the effluents during wastewater irrigation is one major route of infection. STH egg concentrations in the final effluents from the centralized and DEWATS wastewater treatment plants were consistently higher than the WHO recommended guideline for unrestricted agricultural use (≤ 1 helminth egg/L), whereby the direct reuse of the effluents for agriculture was found to pose a higher risk than the WHO tolerable risk of infection (1 ×10-2 pppy) for farmers and consumers. Annually the use of effluents from the DEWATS plants poses the least risk of infection (1.9 ×10-2 (±2.4×10-4)), which is marginally higher than the WHO tolerable risk value. Well maintained DEWATS plants are more efficient in removing or reducing the concentration of STH eggs in wastewater and therefore pose the least risks of infection compared to centralized wastewater treatment plants. Consumers of vegetables from these farms are also at considerable risks of STH infections. Probabilistic assessment of the STH infection risks showed that farmers applying sludge from Durban and Dakar without adequate protective measures had risks of infections higher than the WHO tolerable risks figure (1×10-2 pppy). Based on the estimated risks of infection after decay, exposure to farm soil after 40-50 days of sludge application may reduce the risks of infection to levels lower the WHO tolerable risks value. However, this may not be practical due to the need for farmers to attend to their crops frequently. Incorporation of the decay of the eggs into the risks assessment also indicated that, using lettuce as a representative vegetable, harvesting of vegetables in Dakar could be done after 40 days of sludge application to reduce the risks of infection to the WHO tolerable value but in Durban harvesting after 30 days ensures that consumers are protected. Therefore, to protect both the farmers and consumers exposed to STH eggs through wastewater/sludge reuse in agriculture the implementation of the WHO multi-barrier approach to risk reduction is required. Risks of STH infections could be directly estimated using epidemiological approaches. By using this approach, the concept of STH infection risks for farmers using wastewater was assessed through direct measurements of the concentration of STHs both in wastewater used for irrigation and the farm soil, as well as the actual load of STHs ova in the stool of farmers and their family members. In Kumasi, Ghana, wastewater used for irrigation of vegetables and the farm soil contained high concentration of STH eggs. There was positive correlation between STH concentrations in the wastewater/soil and STH eggs load in stool of the exposed farmers. Stool analysis after 3 months, following deworming, showed a fast re-infection rate. Farmers exposed to the wastewater were three times more likely as compared to the control group of non-farmers to be infected with Ascaris spp (OR = 3.9, 95% CI, 1.15-13.86) and hookworm (OR = 3.07, 95% CI, 0.87-10.82). These risks of infection were higher in the rainy season than the dry season. This corresponds to a higher egg concentration in wastewater used for irrigation during this period. This indicates a relationship between STH infection and egg concertation in the environment. This study therefore contributes to the evidence-based conclusion that wastewater irrigation contributes to a higher incidence of STHs infection for farmers. In conclusion, this thesis therefore presents a new revised method that can be used to determine the STH egg concentration in different environmental samples. The development of this method also provides an opportunity to comparatively assess the STH egg reduction/removal efficiency of the more commonly used centralized wastewater treatment plants and DEWATS plants. The accurate quantification of viable STH eggs provide inputs for the probabilistic assessment of STH infection risks for different populations exposed to effluents from these two wastewater treatment approaches. This assessment of risks provides a public health perspective to the wastewater treatment. Additionally, it was concluded with the used of this method that drying of sludge for 60 days in Durban or Dakar does not produce sludge of good quality for agricultural application. This was confirmed by the estimates of STH infection risks determined using quantitative microbial risks assessment. This thesis therefore shows the importance of accurate quantification of STH eggs in the determination of infection risks either though QMRA or epidemiological approache / D

Thesis (Ph. D.)--University of Tennessee, Knoxville, 2008. / Title from title page screen (viewed on Mar. 10, 2009). Thesis advisor: Kurt H. Lamour. Vita. Includes bibliographical references.

There is significant concern about the accumulation of potentially toxic elements (PTEs) in soils because of both direct and indirect impacts on human and ecosystem health. Knowledge of the fate and distribution of such contamination can lead to an effective assessment of the hazards to soil biota and the need for protective or mitigation activities. This is a particular challenge due to the heterogeneity of the soil matrix and complexity of the processes that determine PTE availability to soil biota. While whole-cell bacterial biosensors have been proposed as tools in enabling greater confidence in addressing such biological and chemical interfaces their genuine value remains to be realised. The underpinning objective of this work was to link the response of microbial biosensors to detailed chemical analysis and to relate the dose response sensitivity to other biological measurements. To better understand the phenomena of PTE bioavailability, the study considered changes in toxicity within the context of ion competition in both freshly amended and historically impacted soils. The interaction of test bacteria with both free (soil pore water) and sorbed (solid phase) fractions of the target analytes (copper, nickel and zinc) has enabled a better estimation of bioavailability/toxicity of PTEs in soils. In comparison to other assays, the responses of the microbial sensor to Cu, Ni and Zn highlighted its relative sensitivity to PTE contamination. The use of luminescence marked microbial sensors complements the performance of rigorous analytical soil chemistry approaches. Their value in soil pollution should be considered a technique that should be interpreted alongside chemical analysis rather than an alternative as their performance in complex environmental matrixes is yet to be validated.

The aim of this thesis was to gain an understanding of the molecular and ecological basis for the biological control of Pythium by fluorescent pseudomonads. A fluorescent pseudomonad biocontrol agent (BCA), Pseudomonas fluorescens 54/96, identified as a potential candidate for commercial development, was analysed together with transposon induced mutants in a variety of assays for anti-fungal activity (Chapter 2). It was revealed that 54/96 had a fungistatic effect generated by a number of different mechanisms, which included nutrient competition and antibiosis. The synecology of this organism with Pythium was then compared to a similar organism (P. fluorescens SBW25) demonstrating a similar degree of anti-fungal activity (Chapter 3). The similarity of the population dynamics of these two strains prompted an examination of the genetic basis for the anti-fungal activity of the second strain, with the intention of comparing with 54/96 (Chapter 4). Again this revealed a multifactorial mode of action of SBW25 against Pythium. Whilst some mutants with reduced anti-fungal activity were deficient in growth on seed exudate others were unaffected, but the mechanisms appeared to be different to those utilized by 54/96. The comparison of strains was expanded to a larger collection of pseudomonad BCAs which were contrasted by a number of phenotypic and genotypic methods (Chapter 5). Various markers were identified which showed commonality within the different classes of BCA, the most useful of which was cyclopropanated fatty acids. These may prove to be a useful marker when screening for new pseudomonad BCAs. It was concluded that a greater understanding of the molecular, physiological and ecological basis of anti-fungal activity of bacterial will lead to the development of biocontrol strategies with improved efficacy.

Thesis (Ph. D.)--University of Adelaide, Dept. of Crop Protection, 1996. / Includes bibliographical references (leaves 207-220).

Thesis(Ph.D.)--University of Western Australia, 2006.

Thesis (M.S.) in Plant, Soil and Environmental Sciences--University of Maine, 2008. / Includes vita. Includes bibliographical references (leaves 88-93).