Soybean growers in the southern United States commonly deal with disease issues that can limit yield potential. As a result, growers have adopted prophylactic fungicide application targeted at specific growth stages as a standard management practice. However, yield responses to prophylactic applications have become sporadic over time. One major change that has occurred is adoption of cultivars with improved disease tolerances. Therefore, the objective of this research was to evaluate yield response of cultivars representing multiple genetic backgrounds to fungicides applied prophylactically at the R4 soybean growth stage. Experiments were conducted in 2020 and 2021 to determine the yield impact of prophylactic fungicide applications in MG IV or V cultivars across different planting dates and multiple soil textures. A yield increase was observed for soybean treated with a fungicide compared to nontreated soybean. However, a yield response between nontreated and treated soybean within cultivar was not observed, indicating that differing cultivar disease tolerances may not influence soybean yield response to prophylactic fungicide application.

Strawberry is an economically important crop in California, with an estimated value of $2.68 billion in 2023. In California strawberry production, mitigation of low plant health and yield often focuses on major soilborne pathogens, while the contribution of minor pathogens and abiotic disorders to production shortcomings are often overlooked. The objectives of the first project in this thesis are to determine the pathogenicity of multiple minor pathogens and quantify other biotic and abiotic factors that can reduce plant health such as viruses and soil salinity. Two pathogens of the black root rot complex, Pythium spp. and Rhizoctonia spp. as well as Neopestalotiopsis rosae are included in this study. The pathogens were identified using ITS DNA sequencing and evaluated for optimal colony growth temperatures. Over the course of two trials, Koch's postulates of P. ultimum and P. irregulare isolates were confirmed for pathogenicity of strawberry roots and crowns, Rhizoctonia spp. isolates were confirmed over 2 trials for pathogenicity of strawberry roots and crowns, and Neopestalotiopsis rosae isolates were confirmed for pathogenicity of strawberry crowns, leaves, and fruit. In 2022 and 2023, 60 root zone soil samples of symptomatic plants that tested negative for major soilborne pathogens were evaluated for electrical conductivity (ECe). Average soil ECe was 1.17 dS/m ranging between 0.18 and 2.45 dS/m, categorizing all samples as non- or slightly-saline except for two which were moderately-saline. Virus testing between 2022 and 2024 diagnosed eight positive samples out of 38 total samples: three in 2022 for Beet pseudo-yellows, two samples in 2023 for Strawberry mild yellow edge and Strawberry polerovirus 1 and one for Strawberry polerovirus 1, and two samples in 2024 for Beet pseudo-yellows. The results from this study suggest that while the tested minor pathogens can infect and reduce strawberry plant health, it is unlikely they are the sole cause of the observed plant mortality from recent diagnostic samples. Additionally, the low to moderate soil salinity levels and infrequent positive virus diagnostics are also not likely the sole cause of observed plant mortality. Future research into these topics could focus on the combination of major and minor pathogens as well as abiotic disorders to decipher how each factor affects plant health. Additionally, Macrophomina phaseolina (MP), an important soilborne pathogen in California strawberry production, was observed to cause 29.7%-52.0% of late-season strawberry mortality in major strawberry growing districts in recent surveys. The objective of the study of this thesis is to assess the efficacy of crop termination and cover cropping on MP suppression to reduce disease incidence of Macrophomina root rot and improve strawberry yield and soil health. Studies were conducted using conventional field soil as a pot trial and a field trial, as well as an organic field trial. The greenhouse pot trial utilized strawberry cultivars Albion and Royal Royce planted in soil collected from a conventional grower field in the Santa Maria district in a randomized complete block design. Treatments included untreated control soil (C), untreated control soil planted with wheat (W) 'Summit 515’ (C+W), soil fumigated with metam potassium (crop termination) planted with wheat (CT+W), and soil fumigated with metam potassium (crop termination) and chloropicrin (flat fumigation) (CT+FF). Plant infection and soil pathogen levels were assessed via plating on semi-selective media and using a pour plate method, respectively. Two repetitions of the trial were conducted. There was no significant soil treatment × cultivar interaction or cultivar effects in trial 1 and 2 for the MP CFU/g soil, but there was a significant soil treatment effect in the MP CFU/g soil of both trials (P = 0.0001). The trial 1 CT+W treatment had the highest-level MP CFU/g soil, which descended in significance to C+W, then C, and then finally CT+FF. In trial 2 CT+W had a significantly higher MP CFU/g soil value than the other treatments, while C and C+W were comparable to each other and both higher than CT+FF. Chemical soil evaluations for mineralizable carbon (MinC) and permanganate oxidizable carbon (POXC) were also performed. Trial 1 C+W had a significantly higher MinC value than CT+FF, while C and CT+W were comparable with both treatments. In the trial 2 MinC soil test the soil treatment × cultivar interaction and the main effects were not statistically significant. In trial 1 and trial 2 POXC soil tests the soil treatment × cultivar interaction and the main effects were not statistically significant. The second year of this study took place in the field and soil samples were collected pre- and post-soil treatments of CT, CT+W, CT+Triticale ‘Pacheco’ (T), CT+W+FF, and CT+T+FF to be evaluated for MP CFU/g soil. T was added as a cover crop treatment to compare a triticale variety to wheat as well as its prevalence as a cover crop in California. There was a significant soil treatment effect (P = 0.02) with post-cover crop wheat treatment having a significantly higher MP CFU/g soil than post-cover crop triticale. The strawberry cultivar Portola was planted after fumigation across all cover crop blocks. Additionally, an organic strawberry field trial compared wheat and triticale cover crops for MP suppression with strawberry cultivars Valiant and Monterey. Soil and plants were tested in the same manner as the pot trial. MP CFU/g soil, MinC, and POXC were not significantly different between soil treatments. Preliminary results suggest single season cover cropping cannot manage high pathogen levels, while crop termination can reduce the pathogen inoculum if the application is timed correctly. The completion of this project will include plant mortality evaluations and microbiome analyses from the field trials. This research aims to help the California strawberry industry by enhancing disease management and reducing fumigant use.

Development of resistant cultivars is the main control measure against sugarcane brown rust caused by Puccinia melanocephala. Durability is uncertain, since the pathogen possesses adaptive ability to overcome host plant resistance. A differential gene expression study utilizing suppressive subtraction hybridization was conducted to improve understanding of brown rust resistance mechanisms in sugarcane. The expression patterns of 11 unigenes representing biosynthetic pathways, defense-related genes, and signaling genes were analyzed in L 99-233, a cultivar exhibiting quantitative resistance, L 01-299, a resistant cultivar with the major resistance gene Bru1, and two susceptible cultivars, Ho 95-988 and L 09-125, at 24 h, 48 h, 72 h, and 1 week after inoculation with P. melanocephala using (semi)quantitative RT-PCR. All genes analyzed for their expression showed message accumulation upon infection in susceptible and resistant cultivars, but the maintenance of high amounts of mRNAs of the genes for a prolonged time period appeared to be the most important factor contributing to brown rust resistance. Differences in the time-course of gene expression were detected between L 01-299 and L 99-233 suggesting variable mechanisms for resistance between the cultivars. Molecular markers were used to screen the World Collection of Sugarcane and Related Grasses (WCSRG) for Bru1 to determine its distribution and frequency in Saccharum species and related genera. A total of 1,282 clones were screened. Bru1 was distributed across the Saccharum complex, but the frequency varied among species. Bru1was more prevalent in S. robustum clones (59.1%), whereas it occurred in low frequency and exhibited the highest level of variability in clones of S. spontaneum (18.8%). Bru1 frequency was highest in the two secondary cultivated species, S. barberi (79.3%) and S. sinense (71.8%). The frequency of Bru1 detection was 26.4% and 21.0% for S. officinarum and interspecific hybrid clones, respectively. The characterization of the WCSRG for Bru1 distribution and prevalence will complement efforts to characterize diversity in the Saccharum complex for the expected expanded use of marker-assisted selection in the future. Selection for quantitative resistance in combination with Bru1 could allow breeding programs to develop sugarcane cultivars with effective and durable resistance against brown rust.

Common bean (Phaseolus vulgaris) is the most important legume for direct human consumption. Common bean originated and was domesticated in the Americas but now is grown worldwide. As in the case of other crops, common bean can be infected with acute and persistent plant viruses. A modified dsRNA extraction method was developed and used in this study. The method was fast, economic, versatile, and required relatively small amounts of desiccated plant tissue. The method was successfully used to extract dsRNAs from plants infected with RNA plant viruses and to investigate the occurrence of two endornaviruses, Phaseolus vulgaris endornavirus 1 (PvEV1) and Phaseolus vulgaris endornavirus 2 (PvEV2), in breeding lines, cultivars, landraces, and wild genotypes of common bean from the two centers of common bean domestication: Mesoamerica and the Andes. The two endornaviruses were detected in many common bean genotypes of Mesoamerican origin but rarely in genotypes of Andean origin. A comparative study of morphological and physiological characteristics between two common bean lines of the cultivar Black Turtle Soup (BTS); one infected with PvEV1 and PvEV2 (BTS+) and the other endornavirus-free (BTS-) was conducted. Morphological differences between the two lines were not observed. However, the study revealed that common bean endornaviruses may promote seed germination, pod length, and carotenoid content. Nevertheless, endornaviruses were associated with lower chlorophyll content. When interactions studies were conducted between PvEV1 and PvEV2 and three acute viruses, synergistic effects were obtained. Quantitative RT-PCR results supported a synergism between PvEV1 and Sunn-hemp mosaic virus. More research should be conducted to determine the type of symbiotic interaction that exists between common bean and endornaviruses.

<p> Soap Lake, located in Washington State, was the subject of an NSF funded Microbial Observatory and is a naturally occurring saline and alkaline lake. Several organisms inhabiting this lake have been identified as producers of siderophores that are unique in structure. Two isolates SL01 &amp; SL28 were the focus of this study of siderophore production, structure elucidation and vesicle self-assembly. Bacterial isolates, enriched from Soap Lake sediment and water samples, were screened for siderophore production. Siderophore production was confirmed through the chrome azurol S (CAS) agar plate method. Isolates SL01 and SL28 were found to produce relatively high concentrations of siderophores in liquid medium. Extraction was performed by the methanol/water protocol in Varian cartridges and siderophore purification was done on HPLC with a 0-70% acetonitrile gradient. Lyophilization or <i> in vacuo</i> evaporation followed in order to store siderophores. Siderophore structure was determined using liquid chromatography and tandem mass spectrometry (LC/MS/MS) with fatty acid methyl ester (FAME) analysis. Vesicle self-assembly studies were performed using dynamic light scattering (DLS) and epifluorescence microscopy (employing cryoembedding and cryosectioning). Three new amphiphilic siderophore families (two from SL01 and one from SL28) were produced by the bacterial isolates, found to be most closely related to <i>Halomonas variablis</i> and <i>Halomonas pantelleriensis</i>, respectively. These siderophores resemble the amphiphilic aquachelin siderophores produced by <i>Halomonas aquamarina</i> strain DS40M3, a marine bacterium. Addition of ferric iron (Fe⁺³) at different equivalents demonstrated vesicle formation and this was confirmed by both DLS and epifluorescence microscopy. Bacteria thriving under saline and alkaline conditions are capable of producing unique siderophores resembling those produced by microbes inhabiting marine environments. Vesicle self-assembly was confirmed quantitatively and qualitatively. Amphiphilic siderophores may have different applications in medical and environmental fields.</p>

Thesis (MScAgric (Plant Pathology))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: Maize is the most important cereal crop produced in southern Africa. Maize producers, processors, and consumers in the region, however, are affected by Fusarium ear rot, a disease caused primarily by the fungal pathogen Fusarium verticillioides that reduces grain quality and potentially contaminates the grain with mycotoxins (fumonisin). Due to the threat of fumonisin to human and animal health, and the economic losses associated with reductions in grain quality, strategies aimed at the prevention of Fusarium ear rot and fumonisin contamination are required. These preventative strategies should be focused on protecting the crop prior to harvest, as damage is known to occur in the field before storage. Chapter 1 provides the reader with a broad overview of maize production in southern Africa, the disease Fusarium ear rot caused by F. verticillioides, and the contamination of grain with fumonisins. Potential disease management practices are summarised, and the role of host resistance and its underlying mechanisms emphasised. Finally, the use of plant breeding and resistance elicitors as methods to enhance host resistance in maize towards Fusarium ear rot and fumonisin contamination are discussed in detail. The planting of maize genotypes with enhanced host resistance potentially offers the most efficient method to reduce Fusarium ear rot and mycotoxin contamination. If plant breeding is to be used to enhance resistance, sources of genetic resistance are required. These sources would ideally be in the form of locally adapted maize genotypes, such as inbred lines. In Chapter 2, maize inbred lines used in local breeding programmes, which are adapted to the production conditions in southern Africa, were evaluated as potential sources of resistance to Fusarium ear rot and fumonisin contamination. If inbred lines with good genetic resistance were to be identified they could be used by breeding programmes to develop commercial maize cultivars with resistance to Fusarium ear rot and fumonisin. Activation of resistance responses in normally susceptible maize genotypes using resistance elicitors could provide a novel management strategy for Fusarium ear rot control, as no commercial cultivars with complete resistance to this disease have been identified in southern Africa. Elicitors have previously been found to induce resistance to plant pathogens, mostly in dicotyledonous crops, but the ability of a range of elicitors to reduce Fusarium ear rot and fumonisin contamination in maize has not been investigated. In Chapter 3, a variety of chemical elicitors that induced resistance in other plant-pathogen systems were selected based on the different defence pathways that they stimulate, and evaluated in field and greenhouse trials. Three commercial maize hybrids were included in the trial, conducted at two different field sites, and the elicitors were tested for their ability to reduce Fusarium ear rot and fumonisin contamination of grain, as well as for their effect on yield. These elicitors could be applied in the field as part of an integrated disease management programme, are environmentally friendly, and would be affordable to commercial producers that produce the majority of maize in South Africa.

Brown rust, caused by Puccinia melanocephala, is an important disease of sugarcane. Breeding for host plant resistance is the primary control measure. Screening for resistance has relied on rating the severity of symptoms caused by natural infection; however, erratic results make this method problematic. A method accomplishing both infection and disease expression under controlled conditions could avoid the problems associated with resistance evaluations under natural infection. Inoculation of seedlings was evaluated to determine whether it could provide accurate resistance ratings in cross appraisal, and inoculation under controlled conditions was evaluated for the potential to accurately determine resistance reactions in clones with known and unknown reactions in comparison to field reactions. Seedlings from crosses between parents with different levels of resistance were inoculated with urediniospores at concentrations ranging from 1 x 103 to 1 x 106 spores per ml. Disease severity was visually assessed at 1 and 2 weeks after inoculation, and resistance ratings were assigned on a modified 1 to 9 scale. Inoculum concentration strongly affected severity and the frequency of resistant progeny in crosses. Brown rust resistance is a heritable trait; however, parental reaction was not a consistent determinant of progeny distribution across resistance rating categories. These results suggest that seedling inoculation may not be suitable for the evaluation of brown rust resistance. Clones were inoculated with 1 x106 spores per ml, and severity was determined as percentage of leaf area occupied by rust lesions by image analysis. Resistance reactions could not be reliably determined for susceptible clones in single inoculations. Controlled conditions inoculation and natural infection results were not correlated. Multiple inoculations under controlled conditions accurately identified resistant and susceptible clones with severe infection resulting from any single inoculation indicating susceptibility. Therefore, controlled conditions inoculation has the potential to be useful in limited studies to characterize parents in a recurrent selection program and for basic studies of resistance to brown rust.

Ralstonia solanacearum (Rs), the causal agent of bacterial wilt of tomato, can cause severe economic losses to tomato growers in Louisiana (LA). Traditional management tactics are ineffective. Resistant cultivars lack durability and have undesirable horticultural traits. In addition, disease resistance is strain specific. For these reasons, many producers in LA have abandoned their fields for tomato production. Although R. solanacearum is endemic in LA, pathogen populations have never been characterized. Tomato samples with bacterial wilt symptoms were collected from Livingston, Tangipahoa, East Baton Rouge and St. Helena parishes in 2015. Bacterial wilt was confirmed using Agdia Inc. Rs-specific immunoflow strips and bacterial streaming test. Fifteen isolates were recovered from five fields and one greenhouse. Strains from LA were characterized and belong to phylotype I and II. Thirty-three percent of the strains were characterized as biovar 1 and 20% as biovar 3. Forty-seven percent of strains were unable to utilize dulcitol, and thus belong to a new biovar 6 classification. None of the isolates belong to a select agent Race 3 biovar 2. A worldwide collection of genetically diverse eggplant, tomato and pepper varieties was screened for resistance to seven Rs strains from LA. One pepper variety (PM702) and one eggplant variety (cv. MM15) were identified as being highly resistant to all seven strains from LA. All of the tomato varieties tested were susceptible to the LA strains. Grafting with resistant rootstock varieties was explored as a sustainable management strategy for bacterial wilt in LA. Three susceptible tomato cultivars (Celebrity, Florida 91 and BHN602) that are commonly produced in LA were grafted onto cvs. MM152 and PM702 resistant rootstocks and onto tomato cv. Hawaii 7996, which is considered a model resistant variety. None of the tomato plants that were grafted to pepper cv. PM702 survived the grafting process. Tomato plants (all varieties) grafted to eggplant (cv. MM152) rootstocks were the most resistant to Rs strains from LA compared to those grafted to Hawaii 7996. This study shows that Rs strains from LA were capable of infecting putatively resistant tomato rootstocks but may be managed by using resistant eggplant rootstocks.

Cercospora leaf blight (CLB) caused in soybean by Cercospora cf. flagellaris is an important disease in Louisiana. It was thought that CLB starts with leaf purpling, and then, as the disease progresses, leaves become blighted. Moreover, it was assumed that accumulation of cercosporin, a red/purple pigment, was the cause of the purple pigmentation in diseased leaves. However, our observations in Louisiana suggested that these two symptoms were not correlated. The first objective of this work was to examine the relationship between purple and blight symptoms as well as their relationships with endophytic colonization of leaves by the fungus and accumulation of cercosporin in soybean leaves. The second objective was to document biochemical changes in purple, blighted and asymptomatic leaves. We demonstrated that purple and blighted leaves were not necessarily correlated, and that cercosporin concentrations in purple leaves were one third that in blighted leaves. Location and cultivar also determined the type of symptoms shown by soybean leaves. This work provides the first report of accumulation of coumestrol (COU) in purple leaves of soybean affected by CLB and demonstrated that COU may be associated with resistance to C. cf. flagellaris via its antioxidant activity. Production of pterocarpin derivatives, a common reaction to biotic and abiotic stresses, may be the cause of purple discoloration of soybean leaves affected by CLB. Results from this work showed that purple CLB leaf symptoms probably are a plant response to low levels of cercosporin produced by the pathogen in its endophytic stage, and blight symptoms are produced when cercosporin production by the pathogen exceeds the plants antioxidant capability.

The AVR4 effector, secreted by Cladosporium fulvum, has been demonstrated to be involved in pathogen virulence. Recent studies further demonstrated that Avr4 is highly conserved among several Cercospora species, indicating a potential important role of this gene in fungal virulence. Therefore, investigation to determine whether this fungal effector gene is present in Cercospora cf. flagellaris (previously known as C. kikuchii), the causal agent of soybean cercospora leaf blight (CLB) disease, and whether it plays any role in CLB disease development, is of great interest. In the present study, the Avr4 gene from C. cf. flagellaris was cloned and mutants lacking the expression of Avr4 were created by homologous recombination to investigate its role in fungal virulence, cercosporin production as well as CLB disease development. The ∆avr4 mutants produced little or no cercosporin in vitro and the mutants also had significantly reduced cercosporin toxin biosynthesis genes expression. Furthermore, ∆avr4 mutants grew faster and were more sensitive to chitinase in vitro than the wild type. Surprisingly, cercosporin could also directly suppress chitinolytic activity in vitro. When inoculated onto detached soybean leaves, these mutants exhibited reduced virulence compared to the wild type and no cercosporin was detected in mutant inoculated soybean leaves. Taken all together, the results suggest that AVR4 may contribute to the virulence of C. cf. flagellaris on soybean through protecting fungal hyphae and regulating cercosporin biosynthesis. Considering the importance of AVR4 in C. cf. flagellaris virulence, we selected a region of this gene for targeted gene suppression through host induced gene silencing (HIGS) to determine whether this can reduce CLB disease in soybean. It was found that HIGS plants carrying the BPMV-Avr4 construct showed less disease symptoms compared to control plants, and the reduction of symptoms was positively correlated with reduction in Avr4 transcript levels and fungal growth. To the best of our knowledge, this is the first study demonstrating the involvement of AVR4 in CLB disease development possibly through regulating cercosporin production as well as protecting fungal hyphae. In addition, this study also showed the potential of using HIGS as a tool to control this important disease of soybean