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Weed resistance to control measures, particularly herbicides, is a growing problem in agriculture. In the case of herbicides, resistance is sometimes connected to genetic changes that directly affect the target site of the herbicide. Other cases are less straightforward where resistance arises without such a clear-cut mechanism. Understanding the genetic and gene regulatory mechanisms that may lead to the rapid evolution of resistance in weedy species is critical to securing our food supply. To study this phenomenon, we exposed young Arabidopsis plants to sublethal levels of one of four weed management stressors, glyphosate herbicide, trifloxysulfuron herbicide, mechanical clipping, and shading. To evaluate responses to these stressors we collected data on gene expression and regulation via epigenetic modification (methylation) and small RNA (sRNA). For all of the treatments except shade, the stress was limited in duration, and the plants were allowed to recover until flowering, to identify changes that persist to reproduction. At flowering, DNA for methylation bisulfite sequencing, RNA, and sRNA were extracted from newly formed rosette leaf tissue. Analyzing the individual datasets revealed many differential responses when compared to the untreated control for gene expression, methylation, and sRNA expression. All three measures showed increases in differential abundance that were unique to each stressor, with very little overlap between stressors. Herbicide treatments tended to exhibit the largest number of significant differential responses, with glyphosate treatment most often associated with the greatest differences and contributing to overlap. To evaluate how large datasets from methylation, gene expression, and sRNA analyses could be connected and mined to link regulatory information with changes in gene expression, the information from each dataset and for each gene was united in a single large matrix and mined with classification algorithms. Although our models were able to differentiate patterns in a set of simulated data, the raw datasets were too noisy for the models to consistently identify differentially expressed genes. However, by focusing on responses at a local level, we identified several genes with differential expression, differential sRNA, and differential methylation. While further studies will be needed to determine whether these epigenetic changes truly influence gene expression at these sites, the changes detected at the treatment level could prime the plants for future incidents of stress, including herbicides. Doctor of Philosophy Growing resistance to herbicides, particularly glyphosate, is one of the many problems facing agriculture. The rapid rise of resistance across herbicide classes has caused some to wonder if there is a mechanism of adaptation that does not involve mutations. Epigenetics is the study of changes in the phenotype that cannot be attributed to changes in the genotype. Typically, studies revolve around two features of the chromosomes: cytosine methylation and histone modifications. The former can influence how proteins interact with DNA, and the latter can influence protein access to DNA. Both can affect each other in self-reinforcing loops. They can affect gene expression, and DNA methylation can be directed by small RNA (sRNA), which can also influence gene expression through other pathways. To study these processes and their role in abiotic stress response, we aimed to analyze sRNA, RNA, and DNA from Arabidopsis thaliana plants under stress. The stresses applied were sublethal doses of the herbicides, glyphosate and trifloxysulfuron, as well as mechanical clipping and shade to represent other weed management stressors. The focus of the project was to analyze these responses individually and together to find epigenetic responses to stresses routinely encountered by weeds. We tested RNA for gene expression changes under our stress conditions and identified many, including some pertaining to DNA methylation regulation. The herbicide treatments were associated with upregulated defense genes and downregulated growth genes. Shade treated plants had many downregulated defense and other stress response genes. We also detected differential methylation and sRNA responses when compared to the control plants. Changes to methylation and sRNA only accounted for about 20% of the variation in gene expression. While attempting to link the epigenetic process of methylation to gene expression, we connected all the data sets and developed computer programs to try to make correlations. While these methods worked on a simulated dataset, we did not detect broad patterns of changes to epigenetic pathways that correlated strongly with gene expression in our experiment's data. There are many factors that can influence gene expression that could create noise that would hinder the algorithms' abilities to detect differentially expressed genes. This does not, however, rule out the possibility of epigenetic influence on gene expression in local contexts. Through scoring the traits of individual genes, we found several that interest us for future studies.

Harvest weed seed control (HWSC) methods concentrate, remove, or destroy weed seeds captured by the combine during harvest. Furthermore, chaff lining uses a chute fitted on the back of a combine to concentrate chaff and weed seed therein into a narrow line. Since chaff amount increases with crop yield, studies aimed to determine how varying crop yield and the associated chaff amount will affect chaff lining control of select weed species, while also examining subsequent crop performance. Objective 1 of this work focused on wheat chaff lining (WCL), and objective 2 studied soybean chaff lining (SCL). Weed species of interest included wild mustard (Sinapis arvensis L.) and Italian ryegrass (Lolium perenne ssp. multiflorum L. Husnot) in WCL and Palmer amaranth (Amaranthus palmeri S.) and common ragweed (Ambrosia artemisiifolia L.) subject to SCL. Each weed species was evaluated in separate experiments, and the SCL experiment included an additional factor of with and without a cereal rye cover crop treatment. Chaff lines mimicked harvest across a range of wheat and soybean yields, with equal weed seed additions (based on existing fecundity and seed shatter phenology data) to each chaff line. A conventional harvest (control) and an outside-the-chaff-line treatment were included, where total fecundity or weed seed rain occurring prior to harvest based on weed species were broadcast respectively. Inhibition of crop and weed emergence as a function of yield and the associated chaff amount was also investigated in the greenhouse. Crop yield across treatments at the field scale (accounts for both chaff lines and outside-the-chaff-line), was not affected in double-crop soybean following WCL and full-season soybean following SCL. Field scale wheat yield in WCL compared to conventional harvest was not different, increased, or decreased in 8, 3, and 1 site-years, respectively. WCL reduced total weed emergence over the combined double-crop soybean and winter wheat growing seasons by 43-54% at the field scale. SCL reduced common ragweed emergence in cereal rye by 64% and 85% in 2 of 3 locations across the soybean growing season. The cover crop did not reduce common ragweed emergence while it was growing, but residual mulch in soybean reduced emergence by 39%. No differences were observed in Palmer amaranth emergence during cereal rye growth, however cereal rye decreased total emergence by 41%. In 6 of 7 Palmer amaranth location-years, SCL decreased field scale weed emergence in soybean by 81%. These results indicate chaff may create an unfavorable environment for weed seed emergence. In both WCL and SCL, greater amounts of chaff caused larger reductions in weed emergence. Objective 3 focused on quantifying the above-ground biomass breakdown of soybean plants into chaff, straw, and seed fractions as they are processed and dispensed by various harvesters. Depending on HWSC system, chaff and straw residues may also be destroyed, removed, or concentrated. Therefore, chaff and straw nutrient composition was analyzed to evaluate the nutrient and economic consequences of HWSC. Our results show average soybean harvest index is 0.57:1. Furthermore, chaff and straw residues equal 13.4% and 68.5% of the seed weight, respectively. Using 5-year average fertilizer prices (2017 – 2021), replacement of N, P, K and S in chaff, straw, and the combination of both residues costs USD 1.58, USD 5.88, and USD 7.46, respectively. Master of Science in Life Sciences In conventional wheat and soybean production, the primary means of weed control is herbicides. If herbicide use is not diversified, a repeated selection pressure drives weeds to evolve resistance to such chemistries. Producers and researchers alike are constantly looking for new ways to combat weeds and herbicide-resistant issues. Originally developed to control nuisance weeds in Australia, harvest weed seed control (HWSC) offers promise in aiding our current herbicide resistance crisis. To further explain HWSC, it is important to know the harvesting mechanism. Many of the row crops including corn, soybean and wheat are harvested using a combine. Combines cut below or tear off plant material to capture the grain or seed which is processed via a threshing system and separated into three fractions: the seed, chaff, and straw. The grain is allocated to a storage bin and eventually removed from the field. In conventional harvest, the remaining crop residue is spread evenly behind the combine across the field to ensure a balanced distribution of organic matter, nutrients, and residue across the field. There is however more than chaff and straw being dispersed. Weeds present in the field at harvest whose seed is retained at crop maturity and at an elevation above the combine header height will inherently end up inside the combine. HWSC are methods intended to capitalize on the combine capturing weed seeds during harvest. Many HWSC approaches to managing weed seed exist, including destruction, removal and concentration of weed seed. Most of this research focuses on only one method of HWSC, chaff lining. Chaff lining utilizes a chute fitted onto the back of the combine and concentrates weed seed and the chaff fraction only into a narrow line behind the combine. Although not directly known, chaff may inhibit future weed emergence within the line due to a mulching effect, intraspecific competition, a greater degree of rotting and increased seed predators. The chute is inexpensive to construct, and there are no additional labor requirements at harvest making it an appealing HWSC option. There is a limited amount of research on chaff lining in North American cropping systems making it a prime HWSC candidate for this thesis. We were curious if chaff lining could benefit wheat and soybean farmers and if crop yield and the associated chaff amount deposited in chaff lines would have any impact on crops planted and weeds placed in lines. Our results indicate chaff lining does not cause field scale yield consequence in double-crop and full-season soybean following wheat and soybean chaff lining, respectively. The effect of wheat chaff lining on wheat field scale yield was variable, but only caused a yield decrease in 1 of 12 experimental locations. Reductions in weed emergence in chaff lining systems compared to conventional indicate chaff may alter the environment to be unfavorable for weed seed emergence. The final objective of this thesis investigates the economic cost of nutrient loss among HWSC systems. Using average fertilizer prices, the cost to apply N, P, K and S concentrated or lost during HWSC in chaff, straw, and the combination of both residues is USD 1.58, USD 5.88, and USD 7.46, respectively.

Cover crops are widely planted in the mid-Atlantic region for their environmental and agronomic benefits, but incomplete or delayed termination can lead to cover crops becoming weeds in the subsequent cash crop, known as volunteers. Studies were conducted from 2020-2022 to evaluate the effect of four commonly grown cover crop species, winter wheat (Triticum aestivum L.), cereal rye (Secale cereal L.), hairy vetch (Vicia villosa Roth), and rapeseed (Brassica napus L.), and four termination timings; 28, 14, 5, and 1 days before corn planting (DBP). Results indicated volunteerism was only an issue with rapeseed. Delaying rapeseed termination resulted in 0, 5, 12, and 22 volunteer plants m-2 at 28, 14, 5, and 1 DBP in corn. In order to manage these rapeseed volunteers, herbicide evaluations were conducted and indicated that atrazine, isoxaflutole, metribuzin, and pyroxasulfone resulted in 92-94% control when applied preemergence. Similarly, atrazine and glyphosate provided 99% rapeseed control and glufosinate resulted in 89% control when applied postemergence. Therefore, volunteers can easily be controlled with commonly used herbicides in corn. Studies were also conducted to evaluate the benefits of these cover crops, which have the potential to overcome the aforementioned risks. Results indicate that hairy vetch produced the most biomass and provided greater control of summer annual grasses, small-seeded broadleaf and large-seeded broadleaf weeds than other cover crops. Biomass accumulation and extent of weed control increased with delaying cover crop termination. Corn yield was greatest following hairy vetch and was least in rapeseed plots. Termination of cover crops 14 DBP planting increased corn yield by 12%; whereas termination at 1 DBP decreased corn yield by 15% as compared to no cover crop-no till plots. Effective termination of cover crops is an important management consideration and information on termination efficiency can help in devising management plans. In order to assist managers by evaluating cover crop termination efficiency, studies were conducted to evaluate selective and non-selective herbicides and a roller crimper for correlating vegetative indices with visible termination efficiency. Among vegetative indices, the Green Leaf Index had the highest Pearson correlation coefficient for wheat (r = -0.79, p =

Peanut (Arachis hypogaea L.) is second major legume crop grown after soybean in the United States, and its productivity is often limited by drought stress. Drought negatively impacts the yield and quality of peanut. Drought stress in peanut causes an annual loss of approximately $520 million in the United States. Improving peanut yield under water deficit conditions is crucial for peanut growers to maintain their profitability in the market. To achieve this, it is essential to either breed or adopt already available drought tolerant cultivars that can produce higher yield under water deficit conditions. Therefore, the objectives of this research were to (1) evaluate five commercially available virginia and runner type peanut cultivars for pod yield stability using multilocation trials by studying G x E interaction across 13 environments including year, location, and irrigation regime. Linn and Binns, AMMI, Shukla, Wricke's, Finlay and Wilkinson stability models were used to determine pod yield stability. Bailey and Sullivan showed higher stability and adaptability across all stability indices whereas Wynne and TUFRunner presented high mean productivity with lesser stability across environments reflecting specific adaptation to just a few environments. Bailey and Sullivan are recommended for sustainable production across the growing region of Virginia and Carolinas. The second objective (2) was identification of drought tolerance related quantitative trait loci (QTL) and genetic markers to facilitate the development of drought tolerant cultivars. Three diverse recombinant inbred line (RIL) populations, derived from crossing lines N05006 x N04074FCT (Pop-1), line N05006 x Phillips, an old virginia-type cultivar (Pop-2), and lines N08086olJCT x PI 585005 (Pop-3) were phenotyped for the Normalized Difference Vegetation Index (NDVI), Canopy Temperature Depression (CTD), SPAD-meter relative chlorophyll content of the leaves (SPAD) and wilting for QTL mapping. Mapping identified 27 minor QTL on eight chromosomes for all physiological characteristics, i.e NDVI, CTD, SPAD and wilting, with logarithmic of odds values ranging from 2.5 to 38.5 and the phenotypic variance explained by these traits from 1.04 to 11.46 %. There were 4 loci on chromosome 2 associated with NDVI in Pop-1 and Pop-3, explaining 1.8 to 10.38% of the phenotypic variation. These genomic regions may be important resources in peanut breeding programs to improve drought tolerance. Further research is needed to increase the marker density in order to fine map the identified QTL and validate markers linked with these regions. Doctor of Philosophy Peanut is a multi million-dollar industry in the United States, but water limitations have a detrimental impact on yield, quality, and grower income. Drought along with aflatoxin contamination are two major challenges faced by U.S peanut industry. Annual losses to peanut caused by drought are around $520 million in the United States. Irrigation can alleviate water shortage in drought prone regions, but around 65% of peanut production in U.S is under rainfed condition, meaning that only rainfall can satisfy peanut crop water requirements. The most feasible and economical solution to peanut growers under these circumstances is to adopt drought tolerant varieties. In this research, our goal was to facilitate breeding drought tolerant cultivars through identification of molecular markers associated with drought tolerance and to identify already available drought tolerant peanut cultivars that could be a game changer for the producers. Therefore, the objectives of my research were to (1) evaluate in multiple environments five commercially available virginia and runner type peanut cultivars for pod yield stability and grade factors. In this study, we considered 13 environments, including 4 years, 4 locations, and 2 water regime. Statistical tools including Linn and Binns, AMMI, Shukla, Wricke's, and Finlay and Wilkinson were used to determine pod yield stability. These stability indices showed that Bailey and Sullivan are more stable and adaptable across different locations in terms of yield, whereas Wynne and TUFRunner presented high mean yield with lesser stability showing specific adaptation to only few environments. Based on stability analysis, Bailey and Sullivan are recommended for sustainable production across different growing region of Virginia and Carolinas. The second objective (2) was to identify drought tolerance related genomic regions using three mapping populations. Phenotyping and genotyping of three diverse recombinant inbred line (RIL) populations, derived from crossing lines N05006 x N04074FCT (Pop-1), lines N05006 x Phillips, an old virginia-type cultivar (Pop-2), and lines N08086olJCT x PI 585005 (Pop-3) were done to find quantitative trait loci (QTL) for drought related traits. These population were phenotyped for the Normalized Difference Vegetation Index (NDVI), Canopy Temperature Depression (CTD), SPAD-meter relative chlorophyll content of the leaves (SPAD) and wilting for QTL mapping. These surrogate traits are related to trait of interest for drought tolerance. NDVI is effective in predicting biomass and yield. Similarly, CTD is associated with transpiration efficiency and carbon dioxide assimilation. Mapping identified 27 minor QTL on eight chromosomes for all physiological characteristics, i.e NDVI, CTD, SPAD and wilting with logarithmic of odds values range from 2.5 to 38.5 and the phenotypic variance explained by these traits ranging from 1.04 to 11.46 %. There were 4 loci on chromosome 2 associated with NDVI in Pop-1 and Pop-3, explaining 1.8 to 10.38% of the phenotypic variation. These genomic regions may be important resources in peanut breeding programs to improve drought tolerance. Further research is needed to increase the marker density in order to fine map the identified QTL and validate markers linked with these regions.

Plants are sessile organisms and have developed varied mechanisms to tolerate stress. One such mechanism is DNA methylation, which plays a vital role within and across generational stress adaptation. To understand the role of DNA methylation in transgenerational stress adaptation, we exposed Arabidopsis thaliana for four generations of sub-lethal doses of glyphosate, trifloxysulfuron, clipping, and shading, which we further classified into the broader categories of stress ecological (shading and clipping) and herbicides (glyphosate and trifloxysulfuron). We analyzed phenotypic and whole-genome bisulfite sequencing data and found that the Arabidopsis phenotype adapts more rapidly to herbicide stress as compared to ecological stresses. DNA methylation changes for glyphosate were minimal after four generations of stress whereas the other three stresses showed dynamic change in the DNA methylation patterns. To understand within generation stress response, Sorghum bicolor was exposed to the same stresses at sub-lethal doses and we analyzed its phenotypic, whole genome bisulfite sequencing, and gene expression responses. Ecological stresses had higher negative impact on S. bicolor plant growth as compared to herbicide stresses. Similarly, we found higher differentially expressed genes for clipping as compared to both herbicides. All four stresses changed the methylome in a unique way; where we found 998 differentially methylated regions (DMR) for trifloxysulfuron, 193 for shading, 141 for clipping and 60 for glyphosate. Out of these DMR's some occurred genic region, which could potentially change gene expression and help plants withstand stress. Overall, DNA methylation can potentially help plants to withstand stress due to their dynamic and specific response to a variety of stresses both transgenerational and within generation. This information to better understand stress adaptation mechanisms in plants and used to develop stress-resilient crops. Doctor of Philosophy Environmental and anthropogenic stresses can negatively impact plant growth and development. Plants can have stress memory through epigenetic changes which helps them withstand stress in future generations. Epigenetics is the field of science where changes on the DNA and not sequence, that can be an addition or deletion of a methyl group, modification of histones, or production of small RNAs. We wanted to understand short and long-term effects of common anthropogenic and ecological stresses on how DNA methylation changes can help plants to withstand stress. We used the model plant Arabidopsis thaliana and the non-model crop/weed Sorghum bicolor. We exposed plants to sub-lethal doses of two herbicides, clipping, and shade stress, at levels high enough to cause significant visible injury but still allowed them to recover and reproduce for a single generation for S. bicolor and four generations for A. thaliana. We found that A. thaliana rapidly more adapts to herbicide stress as compared to ecological stresses. DNA methylation changes for glyphosate were minimal after four generations of stress whereas the other three stresses showed dynamic changes in the DNA methylation patterns. Each stressed impacted S. bicolor phenotype, DNA methylation, and gene expression in unique ways. We found ecological stresses greatly affected the phenotype of the S. bicolor plants as compared to herbicide stresses. Overall, our results showed that stress can cause DNA methylation changes and in transgenerational stress DNA methylation can potentially play a role in stress adaptation. This information could be useful for scientists to further understand stress resilience in plants.

Low temperature injury is among the greatest challenges facing golf courses with ultradwarf bermudagrass (UDB) (Cynodon dactylon (L.) Pers. x C. transvaalensis Burtt-Davy) putting greens in Virginia. This research focused on the impact of turf covers, fungicide programming, core aeration, and trinexapac-ethyl (TE) on UDB cold tolerance, winter quality, and cold de-acclimation (CD). Our results indicate that the use of turf covers significantly increased UDB canopy and soil temperatures when air temperatures were below -3.9°C. Air gaps under covers and the use of double turf covers increased soil and canopy temperatures compared to single covers alone in some instances, but results were inconsistent. Late fall and early winter fungicide applications of chlorothalonil and azoxystrobin improved UDB quality throughout winter dormancy and spring green up. The addition of a pigmented phosphonate significantly improved winter and spring UDB quality. The addition of acibenzolar-S-methyl to fungicide programs did not improve winter UDB quality or spring green up. Summer core aeration programs were evaluated for their impact on spring green up, turfgrass quality, surface firmness, and moisture retention. Spring UDB green up was improved incrementally as surface disruption increased. Treatments with 20%, 15%, and 10% surface disruption produced higher color vs treatments with lower surface disruption. Surface firmness and volumetric water content of UDB were impacted by construction method but were not significantly impacted by core aeration programs. Field research revealed that 'fall only' and 'fall and winter' TE applications improved UDB quality but only 'fall and winter' delayed UDB premature CD in early spring when UDB can be susceptible to low temperature injury. Growth chamber studies evaluated the impact of TE on UDB cold tolerance to -9.4°C x time duration. Regression analysis predicted a 50% mortality exposure point for UDB under TE treatments of 9.84 hours at -9.4°C (r2=0.836) compared to 11.38 hours at -9.4°C (r2=0.671) for non-treated UDB during cold acclimation. Winter and spring scenarios resulted in delayed CD under TE but no differences in cold tolerance when exposed to -9.4°C. Together, these results increase our understanding of the impact of management practices on UDB winter quality, CD, and low temperature injury. Doctor of Philosophy Ultradwarf bermudagrass putting greens are commonly found on golf courses in warm climates. These grasses thrive in heat and humidity but are susceptible to injury or death when exposed to cold temperatures. This research is focused on evaluating management practices that may impact bermudagrass' susceptibility to injury from cold temperature exposure. The cultural practices evaluated include turf covers, fungicide programming, core aeration, and the use of plant growth regulators to manipulate the turfgrasses own self defense mechanisms. Our results show that the use of turf covers significantly increased putting green canopy and soil temperatures when air temperatures were below -3.9°C. Air gaps under covers and the use of double turf covers increased soil and canopy temperatures compared to single covers alone in some instances, but results were inconsistent. Late fall and early winter fungicide applications of commonly-used fungicides improved putting green quality throughout winter dormancy and spring green up. The addition of a green-pigmented phosphonate fungicide significantly improved winter and spring putting green quality. The addition of a plant defense activator, acibenzolar-S-methyl to fungicide programs did not improve winter quality or spring green up. Summer core aeration programs were evaluated for their impact on spring green up, turfgrass quality, surface firmness, and moisture retention. Spring green up was improved incrementally as surface disruption increased. Treatments with 20%, 15%, and 10% surface disruption produced higher color vs treatments with lower surface disruption. Surface firmness and soil moisture content of the putting greens were impacted by construction method but were not significantly impacted by core aeration programs. Field research revealed that 'fall only' and 'fall and winter' plant growth regulator applications improved ultradwarf bermudagrass quality but only 'fall and winter' delayed premature green-up in early spring when the turfgrass can be susceptible to low temperature injury. Growth chamber studies revealed that plants treated with the growth regulator, trinexapac-ethyl were more sensitive to low-temperature exposure than non-treated plants. Together, these results increase our understanding of the impact of management practices on UDB winter quality, CD, and low temperature injury.

Spring dead spot (Ophiosphaerella spp.) (SDS) of bermudagrass (Cynodon dactylon (L.) Pers. x transvaalensis Burtt Davy) is one of the most challenging diseases in the United States transition zone. Six projects were conducted from 2019 to 2022 to better understand the environmental, edaphic, and spatial distribution of SDS epidemics and to examine management strategies for SDS with chemical and cultural practices. A survey of 51 locations provided support of the geographic distribution of Ophiosphaerella species across the Mid-Atlantic United States. Ophiosphaerella herpotricha and O. korrae were isolated from the Mid-Atlantic region, yet O. narmari was not. Cultivars in which parent material originated from the midwestern United States had predominantly O. herpotricha and cultivars in which the parent material originated from the southeastern United States had predominantly O. korrae. In vitro and in situ fungicide efficacy screenings were conducted for O. herpotricha and O. korrae. Additionally, field studies were conducted to optimize fungicide applications and bermudagrass recovery from SDS. Results highlighted that, generally, O. korrae was less sensitive to fungicides than O. herpotricha; the fungicides isofetamid, mefentrifluconazole, penthiopyrad, and pydiflumetofen were generally the most efficacious against SDS; the different fungicide application methods deployed produced mixed results in their effect on fungicide efficacy against SDS with increased efficacy of tebuconazole against SDS with soil surfactant applications and post-application irrigation in certain scenarios; the optimal timing for fungicide applications for SDS was from 13-18°C with tebuconazole and 13-21°C with isofetamid; and nitrogen applications without cultivation practices in the late spring/early summer optimized bermudagrass recovery from SDS. Lastly, a geospatial survey study was conducted to determine the environmental and edaphic factors that influence SDS epidemics. Results were variable with numerous environmental and edaphic factors influencing SDS depending on the year and location; however, soil pH, soil potassium content, and thatch depth were among the most consistent and influential factors on SDS epidemics. Ultimately, these data improve our recommended strategies for successful SDS management. Doctor of Philosophy Spring dead spot is a damaging turfgrass disease that causes aesthetically displeasing symptoms and potential safety and playability concerns for pedestrians and athletes traversing turfgrass surfaces. This disease is caused by three fungal species, and the distribution of these species in the Mid-Atlantic US and the management of spring dead spot epidemics are not well understood. Studies were conducted from 2019 to 2022 to determine the geographic distribution of the species that cause spring dead spot in the Mid-Atlantic, best management strategies for spring dead spot with chemical and cultural practices, and factors in the environment and soil that influence spring dead spot epidemics. Results from the geographic distribution study showed that two of the three fungal species that cause spring dead spot were found in the Mid-Atlantic US, which has major implications on management strategies for the disease. The results from the studies focusing on best management strategies for spring dead spot with chemical and cultural practices highlight that the two fungal species found in the Mid-Atlantic US responded differently to fungicides, few fungicides suppressed the disease to an acceptable level, fungicide application methods provided variable suppression of the disease, optimal timing for fungicide applications was in the fall months when soil temperatures were between 13°C and 18°C, and nitrogen fertilization without cultivation optimized bermudagrass recovery from spring dead spot symptoms. Lastly, the study examining the environmental and soil factors that influence spring dead spot epidemics showed that many factors in the soil and environment influenced spring dead spot epidemics with soil pH, soil potassium content, and thatch depth among the most prevalent. These studies provide turfgrass managers and researchers a better understanding of spring dead spot and allow for more informed management decisions for prevention of and recovery from the disease.

Positive-strand RNA viruses associate with specific organelle membranes of host cells to establish viral replication complexes. The replication protein 1a of brome mosaic virus associates strongly with the nuclear endoplasmic reticulum (ER) membranes, invaginates membranes into the lumen, and recruits various host proteins to establish replication complexes termed spherules. 1a has a strong affinity towards the perinuclear ER membrane, however, the structural features in 1a that dictate its membrane associations and thereby membrane remodeling activities are unclear. This study examined the possible role of an amphipathic α-helix, helix B, in BMV 1a's membrane association. Deletion or single substitution of multiple amino acids of helix B abolished BMV 1a's localization to nuclear ER membranes. Additional reporter-based, gain-of-function assays showed that helix B is sufficient in targeting several soluble proteins to the nuclear ER membranes. Furthermore, we found that the helix B-mediated organelle targeting is a functionally conserved feature among positive-strand RNA viruses of the alphavirus-like superfamily that includes notable human viruses such as Hepatitis E virus and Rubella virus as well as plant viruses such as cucumber mosaic virus and cowpea chlorotic mottle virus. Our results demonstrate a critical role for helix B across members of the alphavirus-like superfamily in anchoring viral replication complexes to the organelle membranes. We anticipate our findings to be a starting point for the development of sophisticated models to use helix B as a novel target for the development of antivirals for positive-strand RNA viruses that belong to the alphavirus-like superfamily. Doctor of Philosophy Among the seven classes of viruses, the positive-strand RNA viruses dominate the domain of viral diseases of the world. Brome mosaic virus (BMV) is a positive-strand RNA virus that infects cereal crops such as wheat, barley, and rice. BMV has a simple genome organization and serves as a suitable model virus to study and characterize positive-strand RNA viruses. The replication of all positive-strand RNA viruses occurs at the organelle membranes of the host. Membrane association of the replication is one of the early steps and a crucial event in the life cycle of positive-strand RNA viruses. One of the proteins produced early on during BMV infection is the replication protein 1a, which is also the master regulator of viral replication; 1a recruits viral factors in addition to hijacking the necessary host factors at the membranous sites to initiate replication. Upon reaching the organelle membranes, 1a induces membrane rearrangements to form viral replication complexes that safeguard the recruited factors from the deleterious effects of the host cell. The structural determinants within 1a that are responsible for such membrane association are unknown. This study explored the potential roles of a short helical motif within the 1a protein for its ability to dictate such site-specific membrane associations. We show here that this helical region is necessary and sufficient for 1a's membrane-binding activity. We also discovered it to be a functionally conserved feature that is responsible for membrane associations in various viruses of the alphavirus-like superfamily that includes some of the notable human viruses such as Hepatitis E virus and Rubella virus in addition to plant viruses such as cucumber mosaic virus and cowpea chlorotic mottle virus.

Plants are surrounded by various threats from the environment such as pathogens, abiotic stresses, and animal attacks. Nutrient content and distribution are essential for plant growth and development as well as plant immunity. Pathogens extract nutrients from host plants to benefit their own growth and reproduction. Sulfate, amino acids, and phosphate are indispensable elements for plant growth, plant nutrition, and plant resistance/susceptibility to disease. However, the role of these nutrients in plant-oomycete interactions is an unexplored area. We developed a hydroponic system to precisely control the nutrients applied to plants. We used Arabidopsis thaliana and Nicotiana benthamiana (N. b) as model plants. Hyaloperonospora arabidopsidis as well as two Phytophthora species, Phytophothora capsici (P. cap) and Phytophothora nicotianae (P. nic) were used as model oomycete pathogens. Hpa is an obligate biotrophic pathogen that obtains nutrients directly from the host plant without causing cell death, while P. cap and P. nic are hemibiotrophic pathogens that display a biotrophic phase followed by a necrotrophic phase where they feed on dead cells. Genomic evidence suggests that these pathogens might obtain nutrients including sulfur in different forms from the host (organic and inorganic respectively). We have optimized the hydroponic system and used Taqman PCR assays and sporangiophore counts to assay the influence of sulfur nutrients on Hpa and P. cap infections. We found that (1) sulfur transporter and metabolism genes play essential roles in plant-oomycete interactions; (2) sulfur is critical components for HR responses against Hpa; (3) low sulfur induces pathogenesis related genes as a systemic acquired response. RNA-seq analysis on Phytophthora-infected Arabidopsis suggested that sulfur transport, assimilation, and metabolism play an important role in plant-oomycete interactions. A second project used RNA-seq analysis on P. nic infected N. b, to identify potential nutrition-related-plant genes that are necessary for full pathogen virulence. RNAi knockdowns of N. b AAP6 (amino acid permease 6) and PHT4 (phosphate transporter 4) genes showed an inhibition of oomycete colonization. These experiments together advance the study on the interplay between nutrient assimilation/metabolism in host plants and oomycete infection which will provide insight into the mechanisms how pathogens intercept nutrients from host. In the long-term, this research could reveal new traits applicable for disease resistance to promote crop and food production. Doctor of Philosophy Plants are surrounded by diverse threats from the environment such as pathogens, abiotic stresses, and animal attacks. Oomycetes are the most destructive group of pathogens, triggering severe food security issues. Phytophthora is an oomycete genus causing serious economic loss. Traditional disease control managements including pesticides, crop rotation and culture practices, are not time- or financially- efficient due to the difficulty in managing oomycete spread and oomycete resistance to chemicals. Thus, new plant genes for resistance to oomycete diseases would have a major impact. Plant nutrients are critically important for plant fitness in every aspect of plant growth and plant immunity. Cellular regulatory networks for sulfur, amino acids, and phosphate assimilation and metabolism networks connect to every aspect of plant activity such as functioning enzymes, formation of chlorophyll, synthesis of proteins, and plant immunity. These nutrients are part of the plant defense system but also can be beneficial nutrients fed to the invading pathogens. Studying how nutrients are involved in the responses to oomycete invasions will provide information to introduce resistance strategies into crops. We utilized oomycete pathogens with different lifestyles to study the interactions and found that some sulfate transporter genes, an amino acid transporter and a phosphate transporter might be manipulated by oomycete to obtain nutrients. Sufficient nutrition is a critical factor for successfully triggering plant immunity but also could be reprogrammed by pathogens for successful infection and development. Our studies gave useful information to understand which plant nutrient genes are important during plant–oomycete interactions. These findings could be useful in identifying or engineering new plant genes to control plant diseases.

Fungi play important roles in various environments. Some of them infect plants and cause economically important diseases. However, many fungal pathogens cause similar symptoms or are even spread asymptomatically, making it difficult to identify them morphologically. Therefore, culture-independent, sequence-based diagnostic methods that can detect and identify fungi independently of the symptoms that they cause are desirable. Whole genome metagenomic sequencing has the potential to enable rapid diagnosis of plant diseases without culturing pathogens and designing pathogen-specific probes. In my study, the MinION nanopore sequencer, a portable single‐molecule sequencing platform developed by Oxford Nanopore Technologies, was employed to detect the fungus Calonectria pseudonaviculata (Cps), the causal agent of the devastating boxwood blight disease of the popular ornamental boxwood (Buxus spp.). Various DNA extraction methods and computational tools were compared. Detection was sensitive with an extremely low false positive rate for most methods. Therefore, metagenomic sequencing is a promising technology that could be implemented in routine diagnostics of fungal diseases. Other fungi may play important roles in the atmosphere because of their ice nucleation activity (INA). INA is the capacity of some particles to induce ice formation above the temperature that pure water freezes (-38°C). Importantly, INPs affect the ratio of ice crystals to liquid droplets in clouds, which in turn affects Earth's radiation balance and the intensity and frequency of precipitation. A few fungal species can produce ice nucleating particles (INPs) that cause ice formation at temperatures ≥ –10°C and they may be present in clouds. Two such fungal genera are Fusarium and Mortierella but little is known about their INPs and the genetic basis of their INA. In my study, F. avenaceum and M. alpina were examined in detail. INPs of both species were characterized and it was found that strains within both species varied in regards to the strength of INA. Whole genome sequencing and comparative genomic studies were then performed to identify putative INA genes. Differential expression analyses at different growth temperatures were also performed. INP properties of the two species shared similarities, both appearing to consist of secreted aggregates larger than 30 kDa. Low temperatures induced INA in both species. Lists of candidate INA genes were identified based on their presence in the strains with the strongest INA and/or induction of their expression at low temperatures and because they either encode secreted proteins or enzymes that produce other molecules known to have INA in other organisms. These genes can now be characterized further to help identify the fungal INA genes in both species. This can be expected to help increase our understanding of the role of fungal INA in the atmosphere. Doctor of Philosophy Fungi are important to life on Earth and play roles in the environments that surround us. On the one hand, fungi can make plants sick and some plant diseases may even cause economic losses to farmers. If the cause of a disease can be identified accurately in an early stage before symptoms develop, disease transmission may be prevented and plants may be protected from disease. However, it is a challenge to find out which fungus causes which disease since symptoms of different fungal diseases look very similar. Typically, we have to wait for plants to become very sick or we have to isolate the fungus that causes a disease to identity it, which may be time-consuming and not lead to precise identification. DNA sequencing technologies have the potential to lead to more sensitive, faster, and more accurate disease diagnosis and, therefore, may help prevent disease outbreaks. In my study, the MinION nanopore sequencer, a small portable device, was used to detect the fungus causing boxwood blight on boxwood. By loading the DNA of unhealthy boxwood on the device, the boxwood blight pathogen was identified within a very short time. Thus, this method is a promising diagnostic method that may be applied to detect other plant fungal diseases as well. On the other hand, fungi may affect Earth's climate by affecting how many water droplets in clouds are frozen, which in turn affects Earth's temperature and how often and how much it rains and snows. Fungi may affect the freezing of water droplets in clouds since some of them have ice nucleation activity (INA), which is the capacity to catalyze ice formation at a higher temperature than the temperature at which pure water freezes (-38°C), and they may be present in clouds. So far, INA has only been found in a few fungi, including the species Fusarium avenaceum and Mortierella alpina, but the mechanism of their INA is poorly understood. In my study, multiple F. avenaceum and M. alpina strains were examined in detail. Two approaches were used. First, strains in each species were compared with each other to find out how strong their INA is. Once it was found that they differed in their strength of INA, their genomes were sequenced and compared to find genes present in the most active strains and missing from the least active strains since it is these genes that may contribute to INA. It was also found that both fungal species had stronger INA when they were grown at lower temperatures. Therefore, the expression of their genes between higher and lower temperatures was compared to find the genes that were more highly expressed at lower temperatures since it is these genes that may cause INA. Based on previous studies, fungal INPs may either consist of secreted proteins or be the products of biosynthetic gene clusters. Therefore, the list of potential genes was reduced by looking for genes encoding either secreted proteins or biosynthetic gene clusters. The list of these potential INA genes will make it easier to identify the INA genes in F. avenaceum and M. alpina and determine the role of fungi in affecting the weather and climate on Earth.

Cuscuta campestris is an obligate parasitic plant that lacks expanded leaves and roots and requires a host to complete its lifecycle. Parasite-host connections occur via an haustorium, a unique organ that acts as a bridge for the exchange of water, nutrients, macromolecules like mRNA, microRNA, and proteins, and microorganisms. Studies of Cuscuta spp. are challenging due to its dependence on the host and other host influences on the parasite. Recent research has shown intriguing aspects of Cuscuta biology like exchange genetic material with its hosts and loss of genes involved in processes such as high photosynthetic rates and defense. We developed new tools and methodologies that allow us to explore C. campestris biology in an unprecedent way. Foremost of these is an axenic method to grow C. campestris on an Artificial Host System (AHS). The AHS allows C. campestris to display its entire life cycle in vitro, including seed production. Using the AHS, we studied haustorial function, determining the role of nutrients and phytohormones on parasite haustorium development and growth, and found genes involved in haustorial function. The AHS allowed us to demonstrate the positive effect of light on C. campestris growth in the absence of a photosynthetic host and to investigate carotenoid- and ABA- related processes in the haustorial regions. We also wanted to understand how C. campestris defenses work independently from a plant host, so we studied the parasite responses to the bacterial epitope flg22 and the bacteria Peudomonas syringe. Our findings indicate that C. campestris is able to sense flg22, but its response differs from those observed in other non-parasite plants. Transcriptomic analysis revealed up-regulation of genes related to biotic and abiotic stresses, and downregulation of genes related to cuticle development. Our study contributes to understanding the C. campestris immune response in the absence of a host plant. Taken together, this research contributes novel methodologies that enable insights into C. campestris biology without the interference of a plant host on the parasite. Doctor of Philosophy Field dodder (Cuscuta campestris) is a parasitic plant that lacks leaves and roots and attacks a wide range of plants, such as tomato and beets. Dodders are not able to carry out full photosynthesis and thus are incapable of producing enough food or obtaining water to survive on their own, so they parasitize other plants. Dodders have developed specialized structures called haustoria that allow them to take resources directly from their hosts. Studying dodder is challenging due to the dependence of the parasite on its host, such that effects of one plant on the other are hard to disentangle. We developed new tools and methodologies that allow us to explore the biology of dodder in unprecedent ways. We developed an Artificial Host System (AHS) that allows the growth and study of dodder without involving a living host plant. Thanks to this new tool, we were able to improve understanding of the function of the haustorium, discover nutrients and growth factors that are indispensable for dodder development, and prove that dodder growth benefits from light. Using the AHS, we compared haustorial regions and shoot tips of dodder to identify genes specific to haustorial function. Additionally, we studied the responses of dodder to bacteria to understand how it reacts to microbial colonization. Our studies contribute with the development of novel methodologies that allow unprecedent discoveries into the biology of dodder. We expect that this work will promote the study of parasitic plant biology.

Amino acids are necessary for protein synthesis and specialized metabolism in plants. Yet very little is known about how plants sense and regulate when and where to allocate amino acids to meet the demand for nitrogen in growing tissues. In particular, while characterized in yeast and mammals, no amino acid sensor has been identified in plants. Amino Acid Permease 1 (AAP1) has been previously characterized and was shown to mediate amino acid uptake from the soil. aap1 knockout plants and several EMS mutants affected in AAP1 sequence display enhanced tolerance to toxic concentrations of amino acids. Yet, two of the corresponding variant proteins appear to be functional transporters, effectively dissociating amino acid transport and phenotype. To understand this apparent discrepancy, I precisely studied AAP1 localization of expression at the plant and cellular level, and in specific tissue types of the root where AAP1 function is required for the tolerance phenotype and the amino acid uptake activity. I showed that AAP1 protein is present in the endoplasmic reticulum of the cortex in wild type plants Yet, its ectopic expression in root tip and phloem increased amino acid uptake, while expression in cortex could not. This and other of my results do not support the current model of AAP1 functioning in amino acid uptake by the root. I propose that the main effect of mutations in AAP1 is a disturbance in amino acid metabolism, possibly triggered by altered amino acid sensing. In this new model, AAP1 would be necessary for sensing amino acid status of cortex cells, possibly in the endoplasmic reticulum, and adjust amino acid metabolic activity and uptake to current availability. In effect, disruption of the sensing function, either by complete loss of AAP1 function (knockout) or by uncoupling the transport and sensing function (EMS mutants), would lead to the various characteristics of the phenotype of the aap1 mutants I observed. My main hypothesis is that AAP1 is a transporter endowed with sensing function, i.e., an amino acid transceptor. Doctor of Philosophy Changing environments create challenges for plants to grow under harsher, nutrient limiting conditions. Nitrogen is an essential nutrient for plant growth, used for the synthesis of amino acids and other nitrogen-containing metabolites. Amino acids are necessary for protein synthesis and other specialized metabolism – being targets for manipulation for improving agronomic traits. Protein content is a complex trait that involves many genes, possibly including amino acid transporters. In addition, the amount of nitrogen needed by and available to the plant increases or decreases depending on the environment conditions. How plants control nitrogen need and use at the molecular level is not well understood. The data presented here challenge a current model and I report how a protein (AAP1) involved in the acquisition of amino acids from the soil provides regulatory control over these processes. . This valuable information is useful for better understanding how plants use nitrogen and more precise breeding methods can be used to improve traits, such as protein content in agronomically important crops.

Annual bluegrass weevil (Listronotus maculicollis Kirby) (Coleoptera: Curculionidae) (ABW) is a major pest of annual bluegrass (Poa annua L.) and creeping bentgrass (Agrostis stolonifera L.) on golf courses in the northeastern United States. The asynchronous life cycle makes managing ABW difficult, putting emphasis on scouting to achieve accurate insecticide timing and acceptable control. Little is known about the biology and management of ABW in Virginia's more temperate climate. Reported cases of ABW resistance to pyrethroids (IRAC Group 3) continues to grow in the northeast, yet no pyrethroid-resistance cases have been reported in Virginia outside of the metropolitan Washington, D.C. For this thesis, I confirmed the widespread distribution of ABW across Virginia with a survey of golf course superintendents. Two golf courses in southwestern Virginia were monitored weekly during the 2019 and 2020 growing seasons to determine the seasonal biology of ABW within this region. These data suggest that overwintering ABW emerge much earlier than described in the northeast, with adult weevil activity beginning in late February or early March. I observed three complete ABW generations, with a potential fourth generation. Soil plugs from the same two golf courses were used to compare the salt floatation and Berlese-Tullgren funnel methods of larval extraction. The methods were highly correlated (R2 = 0.7856), suggesting either method is appropriate for ABW larval extraction. Bioassays conducted on adult ABW from the same two golf courses showed that field rate concentrations of the pyrethroid bifenthrin showed variable mortality ranging from 20% to 80% suggesting the presence of resistance genes in the population. A 100-fold rate of bifenthrin resulted in 100% mortality of ABW, however. Because cross-resistance has been reported among northeastern ABW populations, the common insecticide active ingredients chlorpyrifos (IRAC Group 1B: Organophosphate), trichlorfon (IRAC Group 1B: Organophosphate), λ-cyhalothrin (IRAC Group 3: Pyrethroid), α-cypermethrin (IRAC Group 3: Pyrethroid), imidacloprid (IRAC Group 4A: Neonicotinoid), and spinosad (IRAC Group 5: Spinosyn), were tested on ABW adults. In another bioassay, two organophosphates, trichlorfon and chlorpyrifos, resulted in significantly higher ABW mortality rates than all other labeled insecticides (P < 0.0001). Two other larvicides, spinosad and α-cypermethrin, also exhibited adult control, an important factor to consider for ABW management and preventing pyrethroid-resistance. These results provide valuable insight into the seasonal biology and management of ABW in Virginia and direction for further investigation into these populations. Master of Science in Life Sciences Annual bluegrass weevil (Listronotus maculicollis Kirby) (ABW) is a tiny, but damaging insect pest of cool-season golf course turfgrasses in the northeastern United States. As pest populations have spread southward, ABW has become an emerging pest in Virginia. The objectives of this thesis were to 1) determine the geographic distribution and seasonal biology of ABW in Virginia, 2) compare two methods for extracting ABW larvae from turf cores in order to estimate larval densities, and 3) to assess the susceptibility of ABW to various insecticides commonly used by golf course superintendents. A survey of golf course superintendents in 2019 revealed widespread prevalence of ABW across Virginia, particularly in areas growing predominantly cool-season turfgrasses. Two golf courses in southwestern Virginia were monitored weekly during the growing seasons of 2019 and 2020 to determine the seasonal biology of ABW. Data suggest that overwintering ABW emerge much earlier than reported in the northeast, with adult weevil activity beginning in late February or early March compared to April in the northeast. In addition, I detected three complete ABW generations, with a possible fourth generation occurring. Soil plugs from two golf courses were used to compare a heat extraction method using a Berlese-Tullgren funnel with the traditional salt float method for extraction of ABW larvae. The two methods extracted similar numbers of ABW larvae suggesting that either method is appropriate for ABW larval extraction. Adult ABW from the same two golf courses were collected and subjected to the pyrethroid insecticide bifenthrin at 0.128 kg ai ha-1 (field application rate), 1.28 kg ai ha-1 (10-fold rate), 12.8 kg ai ha-1 (100-fold rate), along with a water control. Results indicated that the field application rate of bifenthrin killed > 50% but

Stagonospora nodorum blotch (SNB), is a major foliar disease of wheat in the mid-Atlantic U.S., is caused by the necrotrophic fungus Parastagonospora nodorum. SNB is managed using cultural practices, resistant varieties, and foliar fungicides. There are increasing trends of severity and incidence of SNB in Virginia and the surrounding mid-Atlantic region, but it is not known if changes in the pathogen population are contributing to this trend. The overall goal of this research was to 1) determine the occurrence of quinone outside inhibitor (QoI) resistance in Virginia populations of P. nodorum infecting wheat, 2) quantify the distribution of G143A mutations conferring fungicide resistance in Virginia populations of P. nodorum, and 3) characterize genetic diversity of P. nodorum populations in Virginia and assess influences of cultivars and environments on population structure and SNB severity. For Objective 1, QoI resistant isolates of P. nodorum were identified from Virginia wheat fields, and this was the first report of QoI resistant P. nodorum in the United States. The G143A substitution in the cytochrome b gene of P. nodorum was associated with reduced QoI sensitivity, and in Objective 2, a state-wide, two-year survey of P. nodorum populations in Virginia determined that the G143A mutation was widespread in the state and among sampled fields the frequency ranged from 5-32% (mean = 19%). For Objective 3, P. nodorum was isolated from five different wheat cultivars across seven locations over two years in Virginia. SNB severity varied by cultivar but greater differences in disease severity were observed among locations and years suggesting environment plays an important role in SNB development. Among the necrotrophic effector (NE) genes examined, SnTox1 was predominant followed by SnTox3, and frequencies of NE genes did not vary by cultivar or location. P. nodorum populations in Virginia had high genetic diversity, but there was no genetic subdivision among locations or wheat cultivars from which individuals were isolated. Results also indicated that the P. nodorum population in Virginia undergoes a mixed mode of reproduction, but sexual reproduction made the greatest contribution to population structure. Overall, this work provides insights into the population biology of P. nodorum in Virginia and information on variability in fungicide sensitivity and cultivar susceptibility to SNB that has implications for the current and future efficacy of fungicides and host resistance for management of SNB. Doctor of Philosophy Wheat (Triticum aestivum L.) is one of the major cereal crops grown worldwide for food, feed, and other products. However, yields of this crop are often limited by fungal diseases including Stagonospora nodorum blotch (SNB) caused by Parastagonospora nodorum. Increasing trends of severity and incidence of SNB may be due to reduced sensitivity of P. nodorum to fungicides or increased virulence of P. nodorum populations on commonly grown cultivars. Fungicides such as quinone outside inhibitors (QoIs) are one of the major classes of fungicides used for disease control and G143A substitution is the most common point mutation associated with complete resistance to QoIs. Therefore, the overall goal of this research was to better understand genotypic and phenotypic variation in Virginia populations of P. nodorum in the context of fungicide sensitivity and susceptibility of wheat cultivars to SNB. The specific objectives were to 1) determine the occurrence of quinone outside inhibitor (QoI) fungicide resistance in Virginia populations of P. nodorum infecting wheat, 2) quantify the distribution of G143A mutations conferring QoI fungicide resistance in Virginia populations of P. nodorum, and 3) characterize genetic diversity of P. nodorum populations in Virginia and assess influences of cultivars and environments on population structure and SNB severity. Results from this research indicate that QoI fungicide resistance occurs in Virginia populations of P. nodorum due to a target site mutation (G143A substitution in the cytochrome b gene), and this mutation is widespread and relatively common in Virginia wheat fields. Based on a multi-year multilocation study, P. nodorum populations were genetically diverse, but there was no genetic subdivision among locations or wheat cultivars. SNB severity varied by location and cultivar, but disease severity was greatest at site-years with moderate springtime temperatures and high rainfall. Overall, this work contributes to a better understanding of P. nodorum populations including the current efficacy of fungicides and host resistance for management of SNB in the region.

Remote sensing techniques are important for detecting disease within the turfgrass canopy. Herein, we look at two such techniques to assess their viability in detecting and isolating turfgrass diseases. First, thermal imagery is used to detect differences in canopy temperature associated with the onset of brown patch infection in tall fescue. Sixty-four newly seeded stands of tall fescue were arranged in a randomized block design with two runs with eight blocks each containing four inoculum concentrations within a greenhouse. Daily measurements were taken of the canopy and ambient temperature with a thermal camera. After five consecutive days differences were detected in canopy – ambient temperature in both runs (p=0.0015), which continued for the remainder of the experiment. Moreover, analysis of true colour imagery during this time yielded no significant differences between groups. A field study comparing canopy temperature of adjacent symptomatic and asymptomatic tall fescue and creeping bentgrass canopies showed differences as well (p

Grapevine crown gall causes significant economic damage in vineyards and nurseries worldwide. Infected vines are not curable due to irreversible genomic transformation by the tumorigenic bacterium Rhizobium vitis. Crown gall results in a gradual decline in yield and vine vigor, then the complete collapse of the vine. R. vitis ARK-1, an antagonistic and non-tumorigenic strain, has been shown to inhibit gall formation when equal cell concentrations of antagonist and tumorigenic strains are co-inoculated in planta. In this study, we tested the efficacy of ARK-1 against higher cell numbers of tumorigenic isolates and evaluated timing and methods of ARK-1 application in in-planta assays in grapevine (Vitis vinifera 'Chardonnay', 'Cabernet Sauvignon', 'Merlot') and tomato (Solanum lycopersicum 'Beefsteak'). ARK-1 significantly (P ≤ 0.01) reduced gall incidence and gall diameter against four times higher tumorigenic bacterial cell numbers. Inoculation of ARK-1 up to 48 hours before or within 6 hours after inoculation with tumorigenic isolates significantly (P ≤ 0.01) reduced gall incidence and gall diameter. Root dipping of grapevine for 24 hours and tomato for 1 hour in the formulated ARK-1 suspension (Kumiai Chemical Industry Co. Ltd., Japan) prior to transplanting in tumorigenic bacteria-infested soil completely inhibited gall formation in the roots. These results are promising and support the development of ARK-1 as a biological control agent to manage grapevine crown gall. Master of Science in Life Sciences Grapevine crown gall is an economically significant disease in vineyards and nurseries worldwide. The pathogen of grapevine crown gall alters plant genome and causes tumor-like gall formation. Infected vines lose yield and vigor and eventually get killed. In this study, we tested the efficacy of a biological control agent called ARK-1. We challenged ARK-1 against higher cell numbers of pathogenic bacteria, evaluated timings, and practical methods of ARK-1 application. We conducted our greenhouse studies in grapevine (Vitis vinifera) cultivars 'Chardonnay', 'Cabernet Sauvignon', 'Merlot' and tomato (Solanum lycopersicum) cultivar 'Beefsteak'. In efficacy test, ARK-1 was effective and reduced 68% gall incidence against a four times higher pathogenic bacteria mixture. In timing assay, the application of ARK-1 up to 48 hours before inoculation of pathogenic bacteria reduced gall formation by more than 95%. Also, ARK-1 was effective when it was applied within 6 hours after inoculation of pathogenic bacteria. Root soaking of grapevine and tomato in the formulated ARK-1 suspension (KUF-1511, Kumiai Chemical Industry Co. Ltd., Japan), prior to transplanting in pathogenic bacteria-infested soil, completely inhibited gall formation in the roots. The promising result of this study is a step forward towards use of ARK-1 as a biological control agent to manage grapevine crown gall disease.

Stem rot of peanut (Arachis hypogaea L.), caused by a soilborne fungus Athelia rolfsii (Curzi) C. C. Tu and Kimbr. (anamorph: Sclerotium rolfsii Sacc.), is one of the most important diseases in peanut production worldwide. Though new varieties with increased partial resistance to this disease have been developed, there is still a need to utilize fungicides for disease control during the growing season. Fungicides with activity against A. rolfsii are available, and several new products have been recently registered for control of stem rot in peanut. However, fungicides are most effective when applied before or during the early stages of infection. Current scouting methods can detect disease once signs or symptoms are present, but to optimize the timing of fungicide applications and protect crop yield, a method for early detection of soilborne diseases is needed. Previous studies have utilized optical sensors combined with machine learning analysis for the early detection of plant diseases, but these studies mainly focused on foliar diseases. Few studies have applied these technologies for the early detection of soilborne diseases in field crops, including peanut. Thus, the overall goal of this research was to integrate sensor technologies, modern data analytic tools, and properties of standard and newly registered fungicides to develop improved management strategies for stem rot control in peanuts. The specific objectives of this work were to 1) characterize the spectral and thermal responses of peanut to infection with A. rolfsii under controlled conditions, 2) identify optimal wavelengths to detect stem rot of peanut using hyperspectral sensor and machine learning, and 3) evaluate the standard and newly registered peanut fungicides with different modes of action for stem rot control in peanuts using a laboratory bioassay. For Objective 1, spectral reflectance and leaf temperature of peanut plants were measured by spectral and thermal sensors in controlled greenhouse experiments. Differences in sensor-based responses between A. rolfsii-infected and non-infected plants were detected 0 to 1 day after observation of foliar disease symptoms. In addition, spectral responses of peanut to the infection of A. rolfsii were more pronounced and consistent than thermal changes as the disease progressed. Objective 2 aimed to identify specific signatures of stem rot from reflectance data collected in Objective 1 utilizing a machine learning approach. Wavelengths around 505, 690, and 884 nm were repeatedly selected by different methods. The top 10 wavelengths identified by the recursive feature selection methods performed as well as all bands for the classification of healthy peanut plants and plants at different stages of disease development. Whereas the first two objectives focused on disease detection, Objective 3 focused on disease control and compared the properties of different fungicides that are labeled for stem rot control in peanut using a laboratory bioassay of detached peanut tissues. All of the foliar-applied fungicides evaluated provided inhibition of A. rolfsii for up to two weeks on plant tissues that received a direct application. Succinate dehydrogenase inhibitors provided less basipetal protection of stem tissues than quinone outside inhibitor or demethylation inhibitor fungicides. Overall, results of this research provide a foundation for developing sensor/drone-based methods that use disease-specific spectral indices for scouting in the field and for making fungicide application recommendations to manage stem rot of peanut and other soilborne diseases. Doctor of Philosophy Plant diseases are a major constraint to crop production worldwide. Developing effective and economical management strategies for these diseases, including selection of proper fungicide chemistries and making timely fungicide application, is dependent on the ability to accurately detect and diagnose their signs and/or symptoms prior to widespread development in a crop. Optical sensors combined with machine learning analysis are promising tools for automated crop disease detection, but research is still needed to optimize and validate methods for the detection of specific plant diseases. The overarching goal of this research was to use the peanut-stem rot plant disease system to identify and evaluate sensor-based technologies and different fungicide chemistries that can be utilized for the management of soilborne plant diseases. The specific objectives of this work were to 1) characterize the temporal progress of spectral and thermal responses of peanut to infection and colonization with Athelia rolfsii, the causal agent of peanut stem rot 2) identify optimal wavelengths to detect stem rot of peanut using hyperspectral sensor and machine learning, and 3) evaluate standard and newly registered peanut fungicides with different modes of action for stem rot control in peanuts using a laboratory bioassay. Results of this work demonstrate that spectral reflectance measurements are able to distinguish between diseased and healthy plants more consistently than thermal measurements. Several wavelengths were identified using machine learning approaches that can accurately differentiate between peanut plants with symptoms of stem rot and non-symptomatic plants. In addition, a new method was developed to select the top-ranked, non-redundant wavelengths with a custom distance. These selected wavelengths performed better than using all wavelengths, providing a basis for designing low-cost optical filters to specifically detect this disease. In the laboratory bioassay evaluation of fungicides, all of the foliar-applied fungicides provided inhibition of A. rolfsii for up to two weeks on leaf tissues that received a direct application. Percent inhibition of A. rolfsii decreased over time, and the activity of all fungicides decreased at a similar rate. Overall, the findings of this research provide a foundation for developing sensor-based methods for disease scouting and making fungicide application recommendations to manage stem rot of peanut and other soilborne diseases.

Biological invasions have been observed throughout the world for centuries, often with major consequences to biodiversity and food security. Tying invasion to species identity and associated traits has led to numerous hypotheses on why, and where, some species are invasive. In recent decades, attention to intraspecific variation among invaders has produced questions about their adaptation to climate, land use, and environmental change. I examined the intraspecific variation of invasive Johnsongrass's (Sorghum halepense (L.) Pers.) seedling stress response, propagule cold tolerance, and large-scale niche dynamics for correlation with populations' climatic and ecotypic (i.e., agricultural vs. non-agricultural) origin. Overall, I found a greater number of home climate effects than ecotypic effects on various traits. Non-agricultural seed from cold climates and agricultural seed from warm climates germinated more and faster, while non-agricultural seedlings showed uniform chlorophyll production regardless of home soil carbon origin, unlike their agricultural counterparts. Neither seedling stress response nor propagule cold tolerance interacted with ecotype identity; however, drought stress varied with population origins' aridity and soil fertility, and seed from warm/humid and cold/dry climates was most germinable. Comparison of seed and rhizome cold tolerance also suggested that the latter is a conserved trait that may be limiting S. halepense poleward range expansion. This physiological limit, an unchanged cold temperature niche boundary between continents and ecotypes, and a narrowed niche following transition to non-agricultural lands all imply low likelihood of spread based on climatic niche shift. Instead, evidence points to range expansion driven primarily by climate change and highlights agriculture's role in facilitating invasibility. This tandem approach to climate and land use as drivers of intraspecific variation is transferable to other taxa and can help refine our conception of and response to invasion in the Anthropocene. Doctor of Philosophy Exotic invasive species are a global problem, threatening biodiversity and biosecurity now and in the future. In the last several decades, ecologists have studied many individual invaders and their traits to understand what drives their spread. More recently, abundant differences in traits between populations within an invasive species have raised questions about humans' role in facilitating invasion through climate change, land use, and other disturbances. I studied the invasive Johnsongrass's (Sorghum halepense (L.) Pers.) response to drought, nutrient limitation, and freezing to detect differences between populations based on their climate and ecotype (agricultural vs. non-agricultural) origin. I also tracked differences in the climates the species occupied across the globe and North America and projected its future distribution under climate change. Overall, I found a greater number of home climate effects than ecotypic effects on various traits. Non-agricultural seed from cold climates and agricultural seed from warm climates germinated the most, while non-agricultural seedlings performed consistently regardless of soil carbon origin, unlike their agricultural counterparts. In addition, drought stress varied with population origins' rainfall and soil fertility, and seed germination favored warm/humid and cold/dry origin. Rhizome (underground stem) cold tolerance appears to be a trait that limits S. halepense poleward range expansion. Along with no change in the coldest climates occupied worldwide and no spread to new climates with transition to non-agricultural lands, this implies that Johnsongrass is unlikely to expand its range without external forces. Instead future range expansion will likely be driven by climate change. This coupled approach to climate and land use affecting invasion is transferable to other species and can help refine both our concepts and response strategies.

Incidence and severity of southern blight caused by Sclerotium rolfsii Sacc. is increasing in the mid-Atlantic region of the U.S. affecting both conventional and organic vegetable production systems. Traditional southern blight management relied on fumigants and fungicides with often inconsistent and uneconomical results. Moreover, with the phase-out of methyl bromide, and the high cost and toxicity of other available fumigants, it is necessary to identify effective and economical southern blight management alternatives. Therefore, the objectives of this research were to 1) evaluate the effect of planting date and potato (Solanum tuberosum L.) cultivar selection on southern blight incidence and tuber yield and quality; 2) compare potato cultivar tuber susceptibility to S. rolfsii in post-harvest settings; and 3) assess the suitability of six cool-season cover crop biomasses and three locally organic materials as carbon sources for anaerobic soil disinfestation (ASD) treatments and their impact on S. rolfsii viability and soil. In a three-year field study, ten commercial potato cultivars and four planting dates per year were evaluated on the Eastern Shore of Virginia. Later potato plantings generally produced greater disease incidence (85-94%) and lower tuber yield (1.8-9.4 Mg ha-1) and quality (47-78%), likely driven by humid, warm weather later in the season that was conducive to disease and detrimental to the potato crop. While no potato cultivar was completely immune to S. rolfsii incidence, cultivar 'Accumulator' consistently had one of the lowest disease incidences (12-71%) and greatest tuber yield among cultivars (6.2-37.3 Mg ha-1), and 'Adirondack Blue' and 'Red Norland' had the overall greatest disease incidence (33-100%). Following harvest of the field experiment, the susceptibility of tubers from the ten cultivars to post-harvest rot caused by S. rolfsii was compared in a laboratory experiment under controlled conditions. 'Atlantic' followed by 'Accumulator' were the least susceptible (3.7-12.6 g of diseased tissue) whereas 'Adirondack Blue', followed by red-skinned potato cultivars had the greatest severity of post-harvest tuber rot (12-17.5 g of diseased tissue). Following three weeks of ASD treatments in growth chamber pot experiments utilizing nine carbon sources incubated at 15/25°C 12/12-h cycle, most carbon sources induced soil anaerobicity (33-90 % iron oxide paint removal), but they did not reduce sclerotial viability compared to nonamended aerobic controls. However, most amended soils undergoing ASD induced greater soil microbial activity (0.7-2.0 % CO2 in vol.) compared to nonamended controls (0.1-0.7 CO2 in vol.), increased soil pH, and some amendments increased soil nitrate accumulation. Overall, results of these studies indicate that southern blight can be effectively managed in potato by coupling early planting dates with cultivar selection, but though promising, additional studies are needed to determine the parameters to effectively decrease S. rolfsii viability with ASD treatments. Doctor of Philosophy The fungal disease southern blight, caused by the soilborne plant pathogen Sclerotium rolfsii Sacc., is increasing in the mid-Atlantic U.S. The increase of southern blight is likely due to a multitude of issues such as climate fluctuations, reduction in the use of fumigants, increased soil organic matter through conservation practices, and cropping systems that favor soilborne pathogens. Traditional management of S. rolfsii relied on pesticides and fumigants, but control was often inconsistent and expensive. Moreover, the use of some soil fumigants has been restricted due to environmental concerns, which has limited the options for the management of plant pathogens present in the soil, and management alternatives are needed. Through a series of field, laboratory, and greenhouse experiments, this study 1) evaluated the effect of planting date and potato (Solanum tuberosum L.) cultivar selection on southern blight incidence and tuber yield and quality; 2) compared potato cultivar tuber susceptibility to S. rolfsii in post-harvest settings; and 3) assessed the suitability of six cool-season cover crop biomasses and three locally-sourced organic materials as carbon sources in anaerobic soil disinfestation (ASD) treatments and their impact on S. rolfsii viability and soil. Results of a three-year field study demonstrated that an integrated approach to the management of southern blight that utilizes moderately resistant potato cultivars such as 'Accumulator' and early (March to early April) planting dates will minimize risk of disease and maximize tuber yield. Following harvest, inoculation of potato tubers under controlled conditions revealed that the cultivar 'Atlantic' followed by 'Accumulator' can avoid greater losses due to less tuber rot, while 'Adirondack Blue' followed by red-skinned potatoes have a greater risk of suffering post-harvest tuber decay from southern blight. The anaerobic soil disinfestation (ASD) treatments examined in this study indicated that cover crops biomass and poultry litter are suitable carbon sources to induce soil anaerobic conditions, but they may not reduce the survival of S. rolfsii propagules if treatment temperatures are below 25°C. However, ASD showed benefits such as increased activity of soil microbial communities compared to nonamended soil, increased soil acidity, and some amendments such as mushroom compost, poultry litter, and Austrian winter pea (Pisum sativum subsp. arvense) can contribute soil nutrients such as nitrate. Overall, this research demonstrates that choosing less susceptible cultivars coupled with early planting dates can minimize the negative impact of southern blight on potato yields and quality, and consequently minimize the use of pesticides as the main option to control southern blight on potato across the mid-Atlantic region. Though promising, additional studies are needed to determine the parameters that optimize ASD as an alternative to manage S. rolfsii with ASD treatments.

Zearalenone (ZEN) is an estrogenic mycotoxin produced by fungi in the genus Fusarium. Fusarium species commonly occur in cereal crops such as corn, wheat, barley, and rice which are used in swine feed. In addition to cereals, co-products of ethanol distillation, brewing, and baking are frequently re-purposed as swine feed. ZEN contamination has been related to severe reproductive issues including pelvic organ prolapse (POP), which describes the displacement of pelvic floor organs from their healthy anatomical positions. A recent rise in the rate of POP in sows highlights a need for more information surrounding the occurrence of ZEN in swine feed and the possible role of ZEN in POP. This research aims to understand the relationship between ZEN contamination in swine feed and swine reproductive health. The primary objectives of this work are to (1) characterize the stress relaxation behavior of vaginal tissue in the swine model, (2) determine how ZEN consumption effects the elasticity of pelvic support tissue. (3) determine where ZEN and a related metabolite accumulate in the swine reproductive tract, (4) evaluate the prevalence of ZEN and related Fusarium mycotoxins in swine feed and ingredients, and (5) track ZEN and related Fusarium mycotoxins throughout production of beer and brewers' spent grains (BSG). Results from this work can help stakeholders to mitigate mycotoxin contamination in swine feed, evaluate swine for ZEN toxicosis, and provide care for animals experiencing POP. Doctor of Philosophy Under certain conditions, some fungi produce harmful toxins (mycotoxins) that can compromise the health of humans and animals. Mycotoxin-producing fungi are common in cereal crops such as corn, wheat, and barley which may be fed to humans or animals. Swine are especially sensitive to mycotoxins, making feed composition important for animal welfare and profitability. Zearalenone (ZEN) is a mycotoxin that behaves like estrogen, a hormone frequently associated with female reproductive health. Pelvic organ prolapse (POP) is a condition, common in women, wherein organs in the pelvic floor such as the vagina, rectum, and uterus, descend from their healthy anatomical position. Swine herds are experiencing higher rates of POP, which may be related to ZEN contamination in feed. The overall goal of this research is to understand the relationship between mycotoxin contamination in feed and swine reproductive health. The specific objectives of this research are to (1) characterize the mechanical properties of vaginal tissue in swine, (2) characterize mechanical properties of pelvic support tissue associated with POP, in swine fed ZEN, (3) determine which reproductive tissues which accumulate ZEN in swine, (4) evaluate the prevalence of ZEN and related mycotoxins in swine feed, and (5) track ZEN and related mycotoxins throughout production of beer and brewers' spent grain, a co-product used in animal feed. This research can help animal producers develop strategies to mitigate mycotoxin contamination in feed and improve veterinary care.

Zoysiagrass (Zoysia spp.) is utilized as a warm-season turfgrass because of its density, visual quality, stress tolerance, and reduced input requirements. Turf managers often exploit winter dormancy in warm-season turfgrass to apply nonselective herbicides such as glyphosate and glufosinate to control winter annual weeds. Although this weed control strategy is common in bermudagrass (Cynodon spp.), it has been less adopted in zoysiagrass due to unexplainable turf injury. Many university extension publications recommend against applying nonselective herbicides to dormant zoysiagrass despite promotional language found in a few peer-reviewed publications and product labels. Previous researchers have used vague terminology such as "applied to dormant zoysiagrass" or "applied prior to zoysiagrass green-up" to describe herbicide application timings. These ambiguous terms have led to confusion since zoysiagrass typically has subcanopy green leaves and stems throughout the winter dormancy period. No research has sought to explain why some turfgrass managers are observing zoysiagrass injury when the literature only offers evidence that these herbicides do not injure dormant zoysiagrass. We sought to explore various herbicides, prevailing temperatures surrounding application, heat unit based application timings, and spray penetration into zoysiagrass canopies as possible contributors to zoysiagrass injury. The results indicated that a wide range of herbicides may be safely used in dormant zoysiagrass. However, as zoysiagrass begins to produce more green leaves, herbicides such as metsulfuron, glyphosate, glufosinate, flumioxazin, and diquat become too injurious. Glufosinate was consistently more injurious regardless of application timing than glyphosate and other herbicides. When temperatures were 10 °C for 7 d following treatment, a delayed effect of glyphosate and glufosinate effect on digitally-assessed green cover loss was noted on zoysiagrass sprigs. In subsequent studies on turf plugs, a 14-d incubation period at 10 °C reduced glyphosate but not glufosinate effects on turf green color reduction. Glyphosate applied at 125, and 200 GDD5C can safely be applied to zoysiagrass while glufosinate applied at the same timings caused inconsistent and often unacceptable zoysiagrass injury in field studies conducted at Blacksburg, VA, Starkville, MS, and Virginia Beach, VA. Zoysiagrass green leaf density was described as a function of accumulated heat units consistently across years and locations but variably by turf mowing height. Turf normalized difference vegetative index was primarily governed by green turf cover but reduced by herbicide treatments, especially when applied at greater than 200 GDD5C. Substantial spray deposition occurred to subcanopy tissue regardless of nozzle type, pressure and height above the zoysiagrass canopy based on spectrophotometric assessment of a colorant admixture. However, increasing nozzle height above the turf canopy and avoiding air induction type nozzles significantly reduced the percentage of green tissue exposed at lower canopy levels. Absorption of radio-labeled glyphosate and glufosinate was up to four times greater when exposed to zoysiagrass stems compared to leaves. Glyphosate translocated more than glufosinate and both herbicides moved more readily from stem to leaf than from leaf to stem Doctor of Philosophy Zoysiagrass (Zoysia spp.) is utilized as a warm-season turfgrass because of its density, visual quality, stress tolerance, and reduced input requirements. Being that zoysiagrass is a warm-season turfgrass, it enters a dormancy period during the winter months. During this period, zoysiagrasses' active growth is halted, and leaves lose their green color and turn a golden-brown color. The winter dormancy period presents turfgrass managers with a unique opportunity to apply nonselective herbicides such as glyphosate and glufosinate to control a broad spectrum of winter annual weeds. Although this weed control strategy is common in bermudagrass (Cynodon spp.), it has been less adopted in zoysiagrass due to turfgrass managers observing unexplainable turfgrass injury. Many university extension publications recommend against applying nonselective herbicides to dormant zoysiagrass despite language found in peer-reviewed publications and product labels suggesting they could be safely applied. Previous researchers have used vague terminology such as "applied to dormant zoysiagrass" or "applied prior to zoysiagrass green-up" to describe herbicide application timings. These terms have led to confusion about when to make these applications since zoysiagrass typically has subcanopy green leaves and stems throughout the winter dormancy period. No research has sought to explain why some turfgrass managers observe zoysiagrass injury when the literature only offers evidence that these herbicides do not injure dormant zoysiagrass. Research projects were designed to explore various herbicides, temperatures surrounding herbicide applications, application timings, and spray penetration into zoysiagrass canopies as possible contributors to zoysiagrass injury. The results indicated that a wide range of herbicides may be safely used in dormant and semidormant zoysiagrass. However, as zoysiagrass begins to produce more green leaves and stems, herbicides such as metsulfuron, glyphosate, glufosinate, flumioxazin, and diquat become too injurious and should be avoided. Across multiple research studies, glufosinate was consistently more injurious regardless of application timing than glyphosate and other herbicides. When temperatures were 10 °C for 7-d following treatment, it delayed zoysaigrass response to glyphosate and glufosinate. In a subsequent study, when temperatures were at 10 °C for a 14-d period, glyphosate and the nontreated reached 50% green cover at the same time, which suggests cold temperatures could mitigate glyphosate injury on zoysiagrass over a 14-d period. The 10 ° temperature only delayed glufosinate injury on zoysiagrass, and no safening was observed. The results also indicated that as temperatures increased, glyphosate and glufosinate rate in which injury was observed increased on the zoysiagrass. Glyphosate applied at 125, and 200 GDD5C can safely be applied to zoysiagrass while glufosinate applied at the same timings caused inconsistent and often unacceptable zoysiagrass injury in field studies conducted at Blacksburg, VA, Starkville, MS, and Virginia Beach, VA. Zoysiagrass injury increased when glyphosate and glufosinate were applied later into the spring when more green leaves were present regardless of location. Accumulated heat units and zoysiagrass green leaf density were closely related, indicating that accumulated heat units could be a useful tool for turfgrass managers to track zoysiagrass spring green-up. Substantial spray deposition was found on subcanopy zoysiagrass leaves and stems regardless of nozzle type, pressure, and height above the zoysiagrass canopy based on recovered colorant at the upper, middle and lower levels of the zoysiagrass canopy. However, avoiding air induction-type nozzles and raising spray height may slightly decrease penetration of spray droplets into a zoysiagrass subcanopy, but a large percentage of droplets still reached the middle and lower canopy layers in this research. Absorption of radio-labeled glyphosate and glufosinate was up to four times greater when applied directly to zoysiagrass stolen compared to leaves. Glyphosate translocated more than glufosinate, and both herbicides moved more readily from stem to leaf than from leaf to stem. These data suggest limiting the number of green zoysiagrass leaves at application would be an effective method to avoid injury zoysiagrass when applying nonselective herbicides

Pythium diseases are common in the greenhouse production of tobacco transplants and can cause up to 70% seedling loss in hydroponic (float-bed) greenhouses. However, the symptoms and consequences of Pythium diseases are often variable among these greenhouses. A tobacco transplant greenhouse survey was conducted in 2017 in order to investigate the sources of this variability, especially the composition and distribution of Pythium communities within greenhouses. The survey revealed twelve Pythium species. Approximately 80% of the surveyed greenhouses harbored Pythium in at least one of four sites within the greenhouse, including the center walkway, weeds, but especially bay water and tobacco seedlings. Pythium dissotocum, followed by P. myriotylum, were the most common species. Pythium myriotylum, P. coloratum, and P. dissotocum were aggressive pathogens that suppressed seed germination and caused root rot, stunting, foliar chlorosis, and death of tobacco seedlings. Pythium aristosporum, P. porphyrae, P. torulosum, P. inflatum, P. irregulare, P. catenulatum, and a different isolate of P. dissotocum, were weak pathogens, causing root symptoms without affecting the upper part of tobacco seedlings. Pythium adhaerens, P. attrantheridium, and P. pectinolyticum did not affect tobacco seeds or seedlings. The consequences of Pythium infection were more likely to be severe when they occurred during seed germination than at seedling emergence, or after plant stem elongation when seedling roots had started to grow into underlying nutrient solutions, depending on the species of Pythium. High and low variation was observed among isolates of P. dissotocum and P. myriotylum, respectively. Pythium myriotylum co-existed with multiple other Pythium or oomycete species in the same environments within tobacco greenhouses, and significant in vitro and/or in vivo interactions between P. myriotylum and some naturally co-existing species were revealed. Pythium porphyrae may have the potential to protect tobacco seeds and seedlings from P. myriotylum infection. Greenhouse Pythium control trials identified ethaboxam, mefenoxam, and copper ionization as potentially promising alternatives to etridiazole for Pythium disease management in tobacco transplant production. The outcomes of this project provide useful new information to better understand the composition, distribution, and diversity of Pythium communities in tobacco transplant greenhouses and to improve Pythium disease management for tobacco transplant production. Doctor of Philosophy Pythium diseases are common in tobacco transplant production and can cause up to 70% seedling losses in hydroponic (float-bed) tobacco transplant greenhouses. However, little is known about the composition and distribution of Pythium communities in tobacco transplant greenhouses. This project began with a tobacco transplant greenhouse survey, in which 12 Pythium species were recovered from center walkways, weeds, greenhouse bay water, and tobacco seedlings. Pythium dissotocum and P. myriotylum were the two types (species) of Pythium most commonly found in the survey. Pythium myriotylum, P. coloratum, and P. dissotocum were aggressive pathogens that suppressed seed germination and caused root rot, stunting, foliar chlorosis, and death of tobacco seedlings. Pythium aristosporum, P. porphyrae, P. torulosum, P. inflatum, P. irregulare, P. catenulatum, and an isolate of P. dissotocum, were weak pathogens causing root symptoms without affecting the upper part of tobacco seedlings. Pythium adhaerens, P. attrantheridium, and P. pectinolyticum did not affect tobacco seeds or seedlings. The symptoms caused by infection by Pythium species differed among host (tobacco) growth stages, except for the most aggressive species, P. myriotylum. High levels of variation were observed among isolates of P. dissotocum, in terms of vegetative growth rate (on V8 agar media) and aggressiveness on tobacco seed and seedlings. Pythium myriotylum was found to co-exist with multiple other Pythium or oomycete species (neighbor isolates) in the same environments within tobacco greenhouses. Significant interactions between P. myriotylum and some neighbor isolates were revealed, and these interactions significantly affect the consequences of P. myriotylum infection of tobacco seeds. Greenhouse Pythium control trials identified two chemical water treatments (ethaboxam and mefenoxam), and a non-chemical water treatment (copper ionization) as potentially promising alternatives to the current standard Pythium control (etridiazole) for Pythium disease management in tobacco transplant production. The outcomes of this project provide useful new information to both better understand the composition, distribution, and diversity of Pythium communities in tobacco transplant greenhouses and to improve Pythium disease management for tobacco transplant production.

Peanut (Arachis hypogaea L.) is an important food crop in the USA and worldwide with high net returns but yield in excess of 4500 kg ha-1 is needed to offset the production costs. Because yield is limited by biotic and abiotic stresses, cultivars with stress tolerance are needed to optimize yield. The U.S. peanut mini-core germplasm collection is a valuable resource that breeders can use to improve stress tolerance in peanut. Phenotyping for plant height, leaf area, and leaf wilting have been used as proxies for the desired tolerance traits. However, proximal data collection, i.e. measurements are taken on individual plants or in the proximity, is slow. Remote data collection and machine learning techniques for analysis offer a high-throughput phenotyping (HTP) alternative to manual measurements that could help breeding for stress tolerance. The objectives of this study were to 1) develop HTP methods using aerial remote sensing; 2) evaluate the mini-core collection in SE Virginia; and 3) perform a detailed physiological analysis on a sub-set of 28 accessions from the mini-core collection under drought stress, i.e. the sub-set was selected based on contrasting differences under drought in three states, Virginia, Texas, and Oklahoma. To address these objectives, replicated experiments were performed in the field at the Tidewater Agricultural Research and Extension Center in Suffolk, VA, in 2017, 2018, and 2019, under rainfed, irrigated, and controlled conditions using rainout shelters to induce drought. Proximal data collection involved physiological, morphological, and yield measurements. Remote data collection was performed aerially and included collection of red-green-blue (RGB) images and canopy reflectance in the visible, near infra-red, and infra-red spectra. This information was used to estimate plant characteristics related to growth and drought tolerance. Under objective 1), we developed HTP for plant height with 85-95% accuracy, LAI with 85-88% accuracy, and wilting with 91-99% accuracy; this was done with significant reduction of time as compared to proximal data collection. Under objectives 2) and 3), we determined that shorter genotypes were more drought tolerant than taller genotypes; and identified CC650 less wilted and with increased carbon assimilation, electron transport, quantum efficiency, and yield than other accessions. Doctor of Philosophy Peanut is a profitable food crop in the USA but has high input costs. Pod yield over 4500 kg ha-1 is required for a profitable production, which is challenging in dry and hot years, and under disease pressure. Varieties tolerant to dry weather conditions (drought) and disease presence are required to sustain production. A collection of 112 peanut varieties is available for researchers to study the mechanisms of tolerance to drought and disease, and identify tolerant varieties to these stresses. Plant characteristics including height, leaf area, and leaf wilting can be used as proxies to estimate stress tolerance and yield, and identify tolerant varieties. How to measure these characteristics is very important. We think that using images collected by a drone and automated analysis by specific computer programs is the easiest, fastest, and most accurate way. Therefore, the objectives of my study were to 1) use drones and cameras to collect images, and computer programs to derive plant characteristics from these images, 2) evaluate the peanut collection to identify varieties with tolerance to drought and disease, and 3) evaluate in depth a sub-set of 28 varieties from this collection under controlled drought conditions to further learn about peanut mechanisms of tolerance to drought and diseases. Field experiments were conducted in 2017, 2018, and 2019, at the Tidewater Agricultural Research and Extension Center in Suffolk, VA. For some tests, we used rainout shelters to mimic drought. We measured plant height, leaf area, color, and wilting, canopy temperature, photosynthesis, and pod yield. From a drone, we collected images in the visible and invisible radiation and, using specific computer programs, estimated plant characteristics with 95% accuracy for height, 88% for leaf area, and 91% for leaf wilting under drought. We concluded that taller varieties were more susceptible to drought than shorter varieties. Peanut varieties CC650 and CC068 had higher end of season yield. The study showed that drought reduced several key mechanisms of photosynthesis including electron transport; and reduced the end of season yield. Variety CC650 performed better under drought than other varieties of the collection.

Host resistance has become a cornerstone of sustainable production of flue-cured tobacco in regions where root-knot nematodes present a threat to growers. Resistance to races 1 and 3 of M. incognita, historically the most significant root-knot nematode threat to tobacco production, is now widely available in commercially available flue-cured tobacco varieties, and is imparted by the gene Rk1. The same gene also provides resistance to race 1 of M. arenaria. The widespread deployment of this resistance has fostered a shift in root-knot nematode population dynamics, as a result of which M. arenaria race 2 has become the predominant root-knot nematode threat in Virginia. A second resistance gene known to impart resistance to M. javanica, Rk2, has also been incorporated into numerous released cultivars in combination with Rk1. This combination has been demonstrated to impart increased resistance to M. incognita and M. javanica relative to either gene alone. In the present work, eleven greenhouse trials conducted from 2017-2019, as well as two trials conducted in 2018 and 2019 on a cooperating farm, investigated the efficacy of this stacked resistance against M. arenaria race 2 and compared the effect of stacking both resistance genes to the effect of either gene alone relative to a susceptible cultivar. We also evaluated how these forms of resistance compare with resistance possessed by a breeding line with resistance reportedly derived from N. repanda to determine if additional, novel sources of resistance to root-knot nematodes previously identified from other species in the genus Nicotiana could play a role in expanding the genetic diversity of germplasm available for the refinement of host resistance in flue-cured tobacco. Additionally, in light of potential new rule making from the FDA mandating reduced nicotine content of cured tobacco leaf, we investigated the relationship between alkaloid (nicotine) content of flue-cured tobacco and root-knot nematode parasitism, while also evaluating nematode parasitism effects on carbohydrate content. Despite considerable variability in our results, particularly under field conditions, our results demonstrate that stacking Rk1 and Rk2 imparts greater resistance to M. arenaria race 2 than either gene alone, but that an entry possessing resistance reportedly derived from N. repanda exhibited significantly greater resistance to root-knot nematodes than the combination of Rk1 and Rk2 based on root galling, and egg mass and egg production. The alkaloid content of flue-cured tobacco did not appear to have an effect on root-knot nematode parasitism under greenhouse or field conditions, but the presence of the nematode did lead to increased accumulation of nicotine in the roots of plants, while translocation of nicotine to leaves was reduced. Conversely, root-knot nematode parasitism was reduced accumulation of carbohydrates in roots, while having no significant effects on leaf carbohydrate content. Ph.D. Root-knot nematodes (Meloidogyne spp.) are microscopic round worms that can cause considerable damage to flue-cured tobacco (Nicotiana tabacum L.), and while not typically responsible for killing plants outright, can reduce the quality of cured tobacco leaf and may predispose plants to a host of other issues, resulting in challenges and economic burdens on growers. Chemicals that effectively control nematodes, which are animals, pose inherent threats to human applicators and may harm the environment in a number of ways, so the use of tobacco varieties that are resistant to root-knot nematodes is increasingly common and essential to sustainable tobacco production. One form of root-knot nematode resistance, called Rk1, has become common and is found in all commercially grown flue-cured tobacco. This form of resistance is effective against 2 ‘races’ of the root-knot nematode M. incognita, which has historically caused tobacco growers the most issues. However, because this resistance is so widely employed, growers have controlled these nematodes, while another species, M. arenaria, has become more prevalent, particularly ‘race’ 2, which is not controlled by Rk1. We know from previous research that another gene, Rk2, provides resistance to some root-knot nematode that Rk1 does not effect, and that combining both genes seems to provide even greater root-knot nematode control than either gene alone. We investigated whether Rk2 is effective at controlling M. arenaria race 2 when it is combined with Rk1 in greenhouse and field experiments. We also investigated how a different, novel type of resistance, which comes from a species of tobacco related to cultivated tobacco, compares with the Rk1/Rk2 resistance in greenhouse trials. Additionally, the FDA has recently suggested that nicotine levels in tobacco leaf should be dramatically reduced to help mitigate adverse human health consequences associated with tobacco consumption. Nicotine may play some role in resistance to root-knot nematode in tobacco, and conversely, root-knot nematodes may impact levels of nicotine, as well as other important chemical constituents of tobacco. We also investigated these questions in greenhouse and field experiments. Our results ultimately demonstrate that combining both Rk1 and Rk2 gives flue-cured tobacco a higher level of resistance to root-knot nematodes than either gene alone, but also suggests that the form of resistance we evaluated from a related Nicotiana species could be even more effective in controlling these nematodes. We observed that the amount of nicotine present in tobacco did not impact nematode parasitism, but that nematode parasitism could lead to lower levels of nicotine in the leaves of plants because the nematodes, which feed on plants roots, cause damage to the plant that interferes in the movement of nicotine from roots to leaves.

The invasive tree-of-heaven, Ailanthus altissima (Miller) Swingle, is widespread and damaging throughout North America. Verticillium wilt disease is emerging as a potentially exciting biological control option for this difficult to control tree. In Virginia, Verticillium nonalfalfae has been confirmed causing significant mortality to A. altissima, while V. dahliae is suspected to be present and causing lower levels of disease. Little else is known regarding these two fungal species in this state. The purpose of this research was to gain a better understanding of how Verticillium wilt impacts A. altissima and its potential as a biological control agent. We first confirmed V. dahliae's presence in Virginia and its pathogenicity to A. altissima using Koch's postulates. We then completed a regional field-inoculation experiment to show that V. nonalfalfae effectively kills and spreads to adjacent A. altissima, regardless of V. dahliae presence or other climate and stand variables. Additionally, we showed that V. dahliae causes lower levels of disease than V. nonalfalfae, and does not spread rapidly. Next, we surveyed all Virginia A. altissima stands known to be naturally infected with V. nonalfalfae to determine whether V. nonalfalfae persists long-term, that it considerably reduces A. altissima numbers, and that its local prevalence may be higher than initially suspected. However, we were unable to infect A. altissima seedlings using soil collected at these infested sites, suggesting that V. nonalfalfae's survival within field soil may be limited. Lastly, using paired A. altissima invaded-uninvaded sites, we found that A. altissima presence is associated with a decreased proportion of native plants and species in the woody and herbaceous understory, but not the germinable seedbank. Furthermore, we found that this impact on the woody understory appears to increase over time, supporting early management actions and helping us predict post-management restoration needs. We conclude that V. nonalfalfae has a high potential of successfully limiting A. altissima throughout Virginia, supporting its registration as a biopesticide. Doctor of Philosophy Commonly called the tree-of-heaven, the nonnative invasive forest-tree Ailanthus altissima, is extensive, damaging, and spreading throughout North America. After finding large areas of declining tree-of-heaven being killed by two different fungal species (Verticillium nonalfalfae and V. dahliae), research has been focused on how to use these fungi to help us manage the tree-of-heaven. In Virginia, V. nonalfalfae has been confirmed killing large numbers of tree-of-heaven, while V. dahliae is suspected to be present in areas with lower levels of decline. The purpose of our research was to gain a better understanding of how these pathogens impact tree-of-heaven and their potential as biological control agents in Virginia. We first confirmed that V. dahliae is present in Virginia and can cause disease on tree-of-heaven. We then inoculated tree-of-heaven stands throughout the state to confirm that V. nonalfalfae effectively kills and spreads to adjacent tree-of-heaven regardless of V. dahliae presence or other climate or site variables. In contrast, we found that V. dahliae only causes low levels of disease and does not spread effectively. Next, we surveyed all known naturally infected V. nonalfalfae sites in Virginia and demonstrated that V. nonalfalfae persists long term within these stands, considerably reducing but not eradicating the tree-of-heaven, and that V. nonalfalfae's local prevalence may be higher than initially suspected. However, when tree-of-heaven seedlings were planted into soil collected from these infested sites, no disease developed, suggesting that V. nonalfalfae's survival within the soil may be limited. Lastly, by looking at tree-of-heaven stands, we found that the tree-of-heaven's presence is associated with a lower percentage of native plants and species in the understory, but not in the seeds present in the soil. In addition, we found that this impact on the woody plants in the understory appears to become more severe over time, supporting managing the tree-of-heaven as soon as possible. We conclude that V. nonalfalfae used as a biological control has a good potential of successfully limiting the tree-of-heaven in Virginia and support its registration as a biopesticide.

Stink bugs, including native brown stink bug (Euschistus servus) and invasive brown marmorated stink bug (Halyomorpha halys), cause damage to a variety of crops including field corn (Zea mays). Frequency and size of stink bug infestations have increased in corn fields in the Mid-Atlantic U.S., and there are growing concerns that these infestations may contribute to reductions in grain quality including increased mycotoxin concentrations. Prior research on native and invasive stink bugs has focused on understanding their biology, the damage they cause, and elucidating effective and economic management strategies. However, few studies examined the potential for stink bugs to facilitate fungal infection and mycotoxin contamination of corn grain. Thus, the objectives of this research were to: 1) assess the relationship between invasive brown marmorated stink bug (H. halys) feeding injuries and fumonisin contamination of field corn in the Mid-Atlantic U.S., 2) determine if stink bugs are a vector for mycotoxigenic Fusarium spp. in corn, and 3) evaluate the efficacy of pesticides for mitigating stink bug feeding injury and associated mycotoxin contamination in field corn. A correlation between H. halys feeding injury and fumonisin concentrations was identified, and the ability of H. halys to increase F. verticillioides infection and fumonisin concentrations in corn was demonstrated in field experiments. Fusarium species including fumonisin-producing F. verticillioides and F. proliferatum were isolated from field-collected stink bugs, and in laboratory experiments, E. servus was able to transmit F. verticillioides to non-infected corn ears after feeding on F. verticillioides-infected corn. In field studies, both fungicide and insecticide reduced stink bug-associated mycotoxin concentrations in corn, but levels of control were inconsistent. Thus, additional tactics that target both the stink bug and Fusarium should be implemented to mitigate risks of mycotoxin contamination in corn. Doctor of Philosophy Native and invasive stink bugs can severely damage crops including field corn. Frequency and size of stink bug infestations in Mid-Atlantic U.S. corn fields have increased, and there is growing concern that this may contribute to reductions in grain quality. Insect feeding injury is a risk factor for fungal infection and mycotoxin contamination in corn. Mycotoxins are toxic chemicals produced by certain fungi that have detrimental health effects on animals including livestock and humans. The relationship between stink bug feeding injuries and mycotoxin contamination in corn grain is not well understood, and management strategies to minimize the risk of mycotoxin contamination in corn need to be identified. The main goal of this research was to characterize interactions between stink bugs and mycotoxin-producing fungi and identify tactics for controlling both the insect pest and pathogen. Specific objectives were to: 1) assess the relationship between invasive brown marmorated stink bug (H. halys) feeding injuries and fumonisin contamination of field corn in the Mid-Atlantic U.S., 2) determine if stink bugs are a vector for mycotoxin-producing Fusarium spp. in corn, and 3) evaluate the efficacy of pesticides for mitigating stink bug feeding injury and associated mycotoxin contamination in field corn. Results from this work indicated that stink bugs have the ability to cause feeding injuries which facilitate invasion of mycotoxin-producing Fusarium species, leading to increases in mycotoxin concentrations in corn grain. Studies also demonstrated that stink bugs can vector Fusarium species during feeding and increase Fusarium infection of corn resulting in subsequent mycotoxin contamination. Field studies indicated that pesticide applications targeting both the stink bugs and mycotoxigenic fungi may be needed to minimize risk of mycotoxin contamination in corn. However, under low pest pressure, application of pesticides is unlikely to be profitable.

Palmer amaranth (Amaranthus palmeri) and common ragweed (Ambrosia artemisiifolia) are two of the most troublesome weeds in soybean. Both weeds possess widespread resistance to glyphosate and acetolactate synthase (ALS) inhibiting herbicides resulting in the use of protoporphyrinogen oxidase- (PPO) inhibitors to control these biotypes, although PPO-resistant biotypes are increasing. New soybean herbicide-resistant trait technologies enable novel herbicide combinations. Combinations of two herbicide sites-of-action (SOA) improved control 19 to 25% and 14 to 19% of Palmer amaranth and common ragweed, respectively, versus using one SOA (mesotrione, dicamba, 2,4-D, or glufosinate alone). Seed production of 5 to 10 cm Palmer amaranth and common ragweed was reduced greater than 76% by fomesafen, auxin (dicamba and 2,4-D), or glufosinate containing treatments. Some weeds survived and set seed even when treated at the proper size. As weed size increased from 10 to 30 cm, control diminished and fecundity increased, underscoring the importance of proper herbicide application timing. Effective preemergence herbicides reduced the number of weeds present at the postemergence application compared to no treatment, reducing the likelihood of herbicide resistance development. Dicamba, 2,4-D, or glufosinate applied alone or auxin + glufosinate combinations reduced Palmer amaranth seed production greater than 95% when applied at first visible female inflorescence; this first report, in addition to previous reports on individual herbicides, indicates this application timing may be useful for soil seed bank management. This research informs mitigation of herbicide resistance spread and development. Master of Science in Life Sciences Over 30 million hectares of soybeans were harvested in 2019 in the United States, totaling over $31 billion in value. Two of the most troublesome weeds in soybean, Palmer amaranth (Amaranthus palmeri) and common ragweed (Ambrosia artemisiifolia) can cause even greater yield reductions in soybean, up to 79 to 95%, respectively. Frequent, exclusive, and repeated use of a single herbicide has led to multiple herbicide-resistance in both of these weeds. Co-applying two effective herbicides reduces the likelihood of resistance development. New soybean varieties have been genetically modified for resistance to herbicides that were previously unusable, allowing new herbicide combinations. Research was established to investigate these herbicide options to control and reduce seed production of Palmer amaranth and common ragweed with the overarching goal of mitigating herbicide resistance, particularly resistance to protoporphyrinogen oxidase (PPO) inhibiting herbicides, which are a critical part of herbicide options in soybean production. Preemergence herbicides are vital tools in herbicide programs, reducing the number of weeds present at a postemergence application and thereby reducing the risk of herbicide resistance development to the postemergence herbicide. PPO herbicides (flumioxazin, sulfentrazone, or fomesafen) applied preemergence reduced Palmer amaranth and common ragweed density at the postemergence application 82 to 89% and 53 to 94%, respectively. The preemergence herbicide used did not affect control four weeks after the postemergence herbicides were applied. Postemergence herbicides were applied targeting three weed heights: 5 to 10 cm (ideal), 10 to 20 cm, and 20 to 30 cm. Control decreased as weed height increased and larger weeds had greater biomass and seed production, underscoring the importance of proper herbicide application timing. The single site-of-action treatments dicamba, 2,4-D, glufosinate, or fomesafen resulted in greater than 85 and 92% morality of 5 to 10 cm Palmer amaranth and common ragweed, respectively. Palmer amaranth and common ragweed control improved by 19 to 25% and 14 to 19%, respectively, when using two herbicide sites-of-action increased versus using one SOA (mesotrione, dicamba, 2,4-D, or glufosinate alone). The use of two herbicide sites of action resulted in maximum biomass reductions, depending on weed height, of 57 to 96% and 73 to 85% for Palmer amaranth and common ragweed, respectively. Dicamba, 2,4-D, glufosinate alone and in combination with fomesafen reduced seed production (relative to the nontreated) of 5 to 10 cm Palmer amaranth and common ragweed greater than 98 and 76%, respectively. Dicamba, 2,4-D, and glufosinate applied alone or auxin (dicamba and 2,4-D) and glufosinate combinations reduced Palmer amaranth seed production greater than 95% when applied at first visible female inflorescence. This indicates that these herbicides may be useful in soil weed seed bank management. This research reinforces the utility of PPO herbicides for preemergence control and their efficacy postemergence when combined with another effective herbicide, a practice known to reduce herbicide resistance development. This research also reinforces the potential for dicamba, 2,4-D, or glufosinate to reduce weed seed production when applied at a delayed timing. Future research should investigate the progeny of these weeds treated with herbicides at a delayed timing to evaluate the potential for this practice to reduce herbicide resistance development.

Plant-associated microbes (collectively the microbiome) are important contributors to plant health. They are known to play roles in increasing yield via improving stress tolerance, promoting growth, and suppressing the activity of plant pathogens. We investigated the wheat seed-head microbiome (phytomicrobiome) as a potential source of resistance to Fusarium head blight (FHB), or scab. FHB is a devastating disease in wheat, and other cereal grains, that causes losses in both quantity, through reduced yield, and quality of grain, through the production of toxins such as Deoxynivalenol. Efforts to combat FHB have focused primarily on breeding cultivars with resistance and applying fungicides. However, new resources for combatting FHB may lie in microbiome-plant interactions. To explore host-microbiome-pathogen interactions, we used field trials to characterize the seed head bacterial community (16S rRNA gene amplicons) across planting locations, host resistance genotypes, varieties, and plant development stages. We identified bacterial amplicon sequence variants (ASVs) present in each sample and then examined ASV community composition based on our variables. Characterizing bacterial relative abundance across samples, we identified 9,063 ASVs. These ASVs clustered according to plant developmental stages or maturity plant, location, and host genotype, but not by variety or maturity group. First, comparing plants at the pre-flowering versus mature grain-head stage, we found that both bacterial community richness and evenness changed significantly. In addition to these developmental changes, we found that bacterial community structure changes across locations, even between locations. Finally, we found that, in the presence of the pathogen, ASVs cluster by host resistance genotype, and that there are important taxonomic groups that are differentially abundant in the presence of the pathogen. Overall, we found that the wheat grain-head microbiome is shaped by environment-host-pathogen interactions, and that these interactions lead to differential abundance of particular community members that may be important in the management of FHB. Master of Science in Life Sciences Plant associated microbes are important contributors to plant health. They are known to play roles in increasing yield via improved stress tolerance, promoting growth, and suppressing plant disease. We investigated the wheat grain-head microbial communities as a source of disease resistance. The disease is called Fusarium Head Blight (FHB) and is caused by Fusarium graminaerum. FHB is a devastating disease in wheat and other cereals, causing losses, through reduced yield and quality through the production of toxins that prohibit use of the grain. To combat FHB, research has focused on developing plants that have resistance and the application of chemical fungicides. However, new resources for combating FHB may lie in the interactions between plants and microbes. This research is focused on identifying microbes that naturally interact with the plant, and how the pathogen, Fusarium, interacts with these beneficial microbes. In field trials, we characterized the microbial community by DNA sequencing technologies across locations, wheat with varying levels of genetic resistance, and wheat developmental stages. First, between the wheat kernel samples of pre-flowering and maturity, we found significant differences in microbial community. Consistent with other studies we found that the largest changes in microbial community composition across different growing locations. Finally, we found an interaction between the grain head microbiome and host resistance state when plants were exposed to the pathogen. Overall, we find that the wheat grain head microbiome is shaped by growing location and through interactions with the plant host and pathogen.

Bacteria are ubiquitously present in every ecosystem on earth. While bacterial communities that reside in specific habitats, called the microbiota, have characteristic compositions, their constituents are exchanged between habitats. To understand the assembly processes and function of a microbial community in an ecosystem, it is thus important to identify its putative sources and sinks. The sources and sinks of the plant leaf microbiome, also called the phyllosphere microbiome, are still under debate. Here, I hypothesized that precipitation is a so far neglected source of the phyllosphere microbiome. Using 16S rRNA amplicon and metagenomic sequencing, I identified the genera Massilia, Sphingomonas, Methylobacterium, Pseudomonas, Acidiphilium, and Pantoea as members of the core rain microbiome in Blacksburg, VA. Further, I used rainwater as a bacterial inoculum to treat tomato plants. I showed that rain-borne bacteria of the genera Chryseobacterium, Enterobacter, Pantoea, Paenibacillus, Duganella, Streptomyces, Massilia, Shinella, Janthinobacterium, Erwinia, and Hyphomicrobium were significantly more abundant in the tomato phyllosphere 7 days post-inoculation, suggesting that these rain-borne bacteria successfully colonized the tomato phyllosphere and had a direct impact on the composition of its microbiome. These results were confirmed by comparing the phyllosphere microbiota of tomato plants grown under greenhouse conditions, and thus never exposed to rain, compared to plants grown outside under environmental conditions, including precipitation. Since a large diversity of bacteria is associated with rain, I also hypothesized that rain-borne bacteria are well adapted to environmental stresses, similar to the stressors microbial biopesticides are exposed to in the field. I thus explored rain as a source of resilient biopesticides to control fire blight, caused by the bacterial pathogen Erwinia amylovora, on apple. In an in-vitro dual culture assay, I identified rain-borne isolates displaying broad-range inhibition against E. amylovora and several other plant pathogens. Two rain-borne isolates, identified as Pantoea agglomerans and P. ananatis, showed the strongest inhibition of E. amylovora. Further experiments showed that these two Pantoea isolates survive under environmental conditions and have a strong protective effect against E. amylovora. However, protection from disease in an orchard was inconsistent, suggesting that the timing of application and formulations must be improved for field applications. Using a UV-mutagenesis screen and whole-genome sequencing, I found that a phenazine antibiotic produced by the P. agglomerans isolate was the likely active molecule that inhibited E. amylovora. Bacterial communities are constantly released as aerosols into the atmosphere from plant, soil, and aquatic sources. When in the atmosphere, bacteria may play crucial roles in geochemical processes, including the formation of precipitation. To understand the potential role of decaying vegetation as a source of atmospheric Ice Nucleation Particles (INPs), I analyzed a historic leaf litter sample collected in 1970 that had maintained Ice Nucleation Activity (INA) for 48 years. A culture-dependent analysis identified the bacterial species Pantoea ananatis and the fungal species Mortierella alpina to have INA and to be present in the leaf litter sample. Further, I determined that both P. ananatis and M. alpina produced heat-sensitive sub-micron INPs that may contribute to atmospheric INPs. The development of new sequencing technologies has facilitated our understanding of microbial community composition, assembly, and function. Most research in bacterial community composition is based on the sequencing of a single region of the 16S rRNA gene. Here, I tested the potential of culture-independent 16S rRNA sequencing of the phyllosphere microbiome for disease diagnosis. I compared the community composition of the microbiome of the aerial parts of cheddar pinks (Dianthus gratianopolitanus) that showed disease symptoms with the microbiome of healthy plants to identify the causative agent. However, I found that the pathogen is probably ubiquitous on cheddar pinks since it was present at similar abundance levels in symptomatic as well as healthy plants. Moreover, the low-resolution of 16S rRNA sequencing did not allow to identify the pathogen at the species or strain level. In summary, in this thesis, I found support for the hypothesis that rain is one of the sources of the phyllosphere microbiome, that rain is a promising source of biopesticides to control plant diseases in the field, that leaf litter is a source of atmospheric INPs, and that 16S rRNA sequencing is not well suited for pathogen identification in support of plant disease diagnosis. Finally, in additional research to which I contributed but that is not included in this thesis, I found that metagenomic sequencing can identify pathogens at the species and strain level and can overcome the limitations of 16S rRNA sequencing. Doctor of Philosophy Bacteria are present in nearly every ecosystem on earth. Bacterial communities that reside in a specific habitat are known as microbiota and have characteristic compositions and functions that directly impact the health of ecosystems. Microbiota associated with plants, the so-called plant microbiota, play a crucial role in plant fitness. Thus, it is important to study the assembly and diversity of plant microbiota and their impact on the ecosystem. The sources of leaf microbiota remain to be elucidated. Here, I have studied the contribution of rainfall to the bacteria that live on and in plant leaves. First, using DNA sequencing, I identified the bacteria present in rainfall in Blacksburg, VA. Then, using rain as bacterial inoculum, I found that some rain-borne bacteria, including members of the genera Pantoea, Massilia, Janthinobacterium, and Enterobacter, are efficient colonizers of tomato leaves. Either absence or low abundance of rain-borne bacteria from tomato leaves never exposed to rainfall confirmed further that bacteria in rain contribute to the assembly of plant leaf microbiota. The identification of all putative sources and sinks of leaf microbiota is important when trying to manipulate them to improve plant health and crop yield. Since I found that rainfall contains many different bacteria, I also studied the potential application of rain-borne bacteria in agriculture. The main limitations of commercial bio-pesticides are their poor survival and limited efficacy in the field. Here, I speculated that rain-borne bacteria are well adapted to environmental stressors and could represent efficient bio-pesticides under field conditions. In fact, I isolated two rain-borne bacteria from the genus Pantoea that strongly inhibited Erwinia amylovora, the causal agent of the fire blight disease of apple, in the laboratory under controlled conditions. However, I observed inconsistent results in a 2-year field trial in an orchard. Using mutagenesis and DNA sequencing, I found the active molecule that likely inhibited E. amylovora, in one of the rain-borne isolates. Finally, the access to newer and cheaper sequencing technologies has recently facilitated the study of bacteria at large scale. Most research of microbiota is based on the sequencing of a single region of one gene, the 16S rRNA gene. Here, I tested the potential of 16S rRNA sequencing of leaf microbiota for disease diagnosis. However, I identified the pathogen in healthy and diseased plants, suggesting its ubiquitous presence. Further, due to the low-resolution of 16S rRNA sequencing, it was impossible to identify the pathogen at the species level. In summary, I found that rain is a source that contributes to leaf microbiota, that rain is a promising source of bio-pesticides to control plant diseases, and that 16S rRNA sequencing is not recommended as a tool to diagnose plant diseases.

Powdery mildew and gray mold diseases have potential to cause significant economic losses to cut flower growers. Information is needed on the effects of these diseases on growers’ returns and the effectiveness of practices to control the diseases. A survey was conducted to examine powdery mildew and gray mold disease management practices on cut flower farms in North Carolina and Virginia. Twenty cut flower farmers completed an online survey and eight of those respondents also participated in an interview. The survey gathered information about the farms, such as its acreage and number of years growing cut flowers, flower genera affected by the diseases, fungicide use and efficiency, and cultural control use and efficiency. The survey and subsequent interviews provided several major findings. First, the floral genera most affected by powdery mildew in North Carolina and Virginia were Zinnia, followed by Dahlia. The genera most affected by gray mold were Ranunculus, followed by Anemone and Dahlia. Second, crop loss due to each disease was mostly at the 0 to 10% level, with one farm losing 91 to 100% of Phlox crops to powdery mildew and another losing 81 to 90% of Tulipa crops to gray mold. Third, 40% of respondents have not used any fungicides for disease management; and for those who have, neem oil was the product they used most frequently. Fourth, all respondents considered sustainability very important when making disease management decisions; this was supported by their extensive use of cultural practices for disease control, in addition to their preference of nonchemical approaches and biorational products over synthetic pesticides. The commonly used cultural controls included, crop rotation, proper watering, disease resistant cultivars, sanitation, proper spacing, and weeding. Findings from this survey will help cut flowers growers in the Mid-Atlantic region to better manage these two important diseases. MALS

While cover crops are actively growing, they compete with winter annual weeds. Studies were conducted to determine the ability of early planted cover crop monocultures and mixtures and a fall applied residual herbicide to compete with winter annual weeds. Cereal rye containing cover crops provided the greatest control of winter weeds in May. Flumioxazin, as a fall applied herbicide, controlled winter weeds in December, but control did not persist until May. Once cover crops are terminated, their residue suppresses summer annual weeds. A greenhouse experiment studying the effects of cereal rye biomass on common ragweed and Palmer amaranth control and light penetration and two field experiments to determine the effects of cereal rye and wheat cover crop biomass terminated with a roller crimper or left standing on summer weed control and light penetration were conducted. For summer weed control, as cover crop biomass increased, weed control increased, light penetration decreased, soil temperature decreased, and soil moisture increased. Standing cover crop residue provided greater control of common ragweed compared to rolled residue until 8400 kg ha⁻¹ of cover crop biomass. As cover crop biomass increased, rolled cover crop residue reduced light penetration compared to standing residue. Wheat cover crop residue increased soil moisture more compared to cereal rye residue. Cover crops must become established to produce adequate biomass to compete with weeds. Herbicide carryover has the potential to reduce cover crop establishment. A study was conducted to evaluate the potential for 30 different residual herbicides applied in the cash crop growing season to carryover to 10 different cover crops. While visible injury was observed, cover crop biomass was similar to the nontreated check in all cases, indicating that herbicide carryover to cover crops is of little concern. Herbicide carryover has few biological effects on establishment of cover crops, under the conditions and herbicides evaluated, that are effective at competing with winter annual weeds and suppressing summer annual weeds. Master of Science in Life Sciences Cover crops are grown after the cash crop has been harvested and terminated before another is planted. They are grown for environmental and agronomic benefits and not harvested. Cover crops improve soil health, reduce erosion, prevent nutrient loss, and control weeds. Cover crops can compete with weeds while they are actively growing. Their residue can create a mulch layer to reduce weed establishment and limit the amount of light reaching weed seed to reduce germination and establishment. As winter cover crops are growing, they compete with winter weeds for sunlight, nutrients, and water. Fall applied herbicides that remain active in the soil are also utilized to control winter weeds in between cash crop growing seasons. Experiments evaluated the ability of cover crop monocultures and cover crop mixtures compared to a fall applied herbicide to compete with winter annual weeds. Monocultures and mixtures of cereal rye, crimson clover, hairy vetch, and forage radish were utilized. Cereal rye containing treatments provided the greatest control of winter weeds in the spring. The fall applied herbicide provided adequate control of winter weeds in December, but control did not persist until the May, indicating that a fall applied herbicide will not provide control of winter weeds from cash crop harvest to the next cash crop planting. As cover crop biomass increases, summer annual weed control increases. Cover crops are usually terminated with herbicide and left standing in Virginia, but the use of a roller crimper lays down residue and creates a mulch layer. Experiments compared the effects of cereal rye and wheat cover crops at different biomass rates terminated with herbicide only (left standing) or a roller crimper and herbicide on summer weed control, light penetrating the cover crop canopy and reaching the soil surface, soil moisture, and soil temperature. As cover crop biomass increased, weed control increased, light reaching the soil surface decreased, soil temperature decreased, and soil moisture increased. Standing cover crop residue provided greater weed control until 8400 kg ha⁻¹ of cover crop biomass was reached. After 8400 kg ha⁻¹ rolled cover crop residue provided greater control, but control from standing and rolled were both greater than 80% compared to the no cover control. Cereal rye intercepted more light than wheat cover crop residue at less than 6000 kg ha⁻¹ of cover crop biomass was achieved. Rolled cover crop residue intercepted more light than standing residue. Established cover crops most produce adequate biomass to effectively control weeds. Herbicides applied during the cash crop growing season to control weeds can remain active in the soil and reduce the establishment of subsequently planted cover crops. Experiments evaluated the potential for different herbicides applied during the cash crop, such as corn, cotton, or soybeans, to remain in the soil at high enough concentrations to injure cover crops commonly utilized in the Mid-Atlantic region. Cover crops utilized were wheat, barley, cereal rye, oats, annual ryegrass, forage radish, Austrian winter pea, crimson clover, hairy vetch, and rapeseed. Results suggest that little potential exists for the herbicides utilized to persist in the soil to injure the five grass cover crop species utilized. There is the potential for some herbicides to injure forage radish, crimson clover, and rapeseed, but no reduction in cover crop biomass was observed, indicating there is little concern for herbicide carryover to cover crops. Residual herbicide carryover has little effect on establishment of cover crops and does not reduce cover crop biomass, under the conditions and herbicides tested in this study. Cover crops can effectively compete with winter weeds, and as cover crop biomass increases, summer annual weed control increases.

Beef cattle production records from two Virginia Department of Corrections herds from 2012- 2019 were analyzed to determine how the cultivar of tall fescue – toxic endophyte-infected or novel endophyte-infected – impacted cattle productivity metrics. Parameters compared were artificial insemination pregnancy rates, calving rates, body weights, and body condition scores of cows and birth weights and weaning weights of calves. The experimental design was a randomized completed block design with year as the replication. Samples of tall fescue were collected from the farm locations and tested for total ergot alkaloid concentrations. The samples from the farm with toxic endophyte-infected tall fescue all contained toxic levels of ergot alkaloids while the samples from the farm with novel endophyte-infected tall fescue indicated that half of the pastures at this location did not have toxic levels of total ergot alkaloids. Artificial insemination pregnancy rates were greater (P = 0.0011) for cows on novel endophyte-infected tall fescue (52 ± 1%) than for cows on toxic endophyte-infected tall fescue (43 ± 1%). Cows on novel endophyte-infected tall fescue had greater calving rates (94 ± 1%) than cows on toxic endophyte-infected tall fescue (89 ± 1%; P = 0.0243). Body condition scores and body weights at pre-breeding were greater for the cows on novel endophyte-infected tall fescue (P=0.0153 and P=0.0456, respectively). Birthweights were reduced for calves on toxic endophyte-infected tall fescue (34 ± 0.9 kg.) than for calves on novel endophyte-infected tall fescue (38 ± 1.4 kg.; P = 0.0257). Calves on novel endophyte-infected tall fescue had greater weaning weights (278 ± 5.9 kg.) than calves on the toxic endophyte-infected tall fescue (254 ± 5.9 kg.; P = 0.0217). Cattle production was improved on the farm with novel endophyte-infected tall fescue compared to cattle production on the farm with only toxic endophyte-infected tall fescue. MALS

All species on Earth occupy limited geographic space. More than a century of observational, experimental, and theoretical work investigating the factors that drive species distributions have demonstrated the importance of the interactions between abiotic, biotic, and demographic factors in determining why species are found where they are. However, it is still unclear when and where these factors interact to set species range limits. Filling the existing knowledge gaps is imperative for the accurate predictions of how species will respond to global change, and particularly for invasive species, many of which are expected to benefit from global change. Here, I sought to investigate the mechanisms that enable, as well as limit, the range expansion of the globally invasive plant Sorghum halepense (L.) Pers. (Johnsongrass). I performed a series of field and laboratory experiments to study population and range dynamics throughout Johnsongrass's North American distribution, and test for the effects of climate, local habitat, and competition on multiple functional traits. I found Johnsongrass consistently demonstrated impressive performance across varying environments, often growing more than 3 m tall, producing hundreds of flowering culms within a single growing season, and maintaining positive population growth rates, even under intense competition with resident weeds. I also found evidence that seed germination has adapted to varying climates encountered during Johnsongrass's range expansion resulting in a shift in the germination temperature niche from warmer to cooler as Johnsongrass spread from warmer climates in the south to more temperate climates in higher latitudes. This shift in the germination temperature niche may have been an important contributing factor in the range expansion of Johnsongrass by enabling the optimization of seed germination in varying climates. On the other hand, results from a field study suggested a possible trade-off between flowering time and growth in populations originating from the range periphery (i.e., range boundary) which may be limiting, or slowing, continued range expansion of Johnsongrass. Together, the outcomes of this work contribute to our understanding of the factors involved in the distribution of species, which is a fundamental goal of Ecology, and essential to accurately predict how invasive species will respond to global change. Doctor of Philosophy Invasive species threaten our natural ecosystems, our agricultural systems, and even our infrastructure, and we spend billions of dollars each year attempting to control them and reduce their negative impacts. Climate change, habitat destruction, and other forms of global change, will benefit many of these species, magnifying their impacts and promoting their invasion into new territories. Because of the damaging effects of invasive species, and the costs to control them, it is imperative that we are able to predict how they will respond to global change so that we can improve plans to reduce their impact and spread. First, we need to understand the processes that promote their invasion across large swaths of land. Just as importantly, we must study the processes that prevent their invasion of certain areas. Here, I investigated some of the processes that have facilitated, as well as hampered, the spread of the invasive plant Johnsongrass. For this work, I used Johnsongrass plants originating from different habitats, including regions where Johnsongrass is highly invasive and those where Johnsongrass is very rare. I found Johnsongrass originating from regions where it is highly invasive were able to grow very large and produce thousands of seeds that were able to germinate under a range of conditions. These traits may have contributed to the invasion success of this species. However, I found a different pattern for plants that originated from regions where Johnsongrass is rare. These plants reached reproductive age earlier and grew smaller across all environmental conditions, potentially due to the less hospitable climates of these range edges. These findings allow us to project into future climate change scenarios, because it is likely that, as temperatures warm, invasive species will be able to invade new regions, where they will impact the work of conservationists, natural resource professionals, agricultural produces, and other land managers.

Herbicide resistant weeds are driving implementation of integrated weed management (IWM). A new tactic to manage weeds is harvest weed seed control (HWSC), which targets weed seeds retained on the plant at crop harvest and either destroys, removes, or concentrates them. Research is limited on the effectiveness of HWSC in US cropping systems. For HWSC to be effective it is important to know when and how many seed are shed from a weed species in relation to crop harvest. Research was conducted to quantify when weed seed are shattered from 6 economically important weed species, four broadleaf (redroot pigweed, common ragweed, common lambsquarters, and common cocklebur) and two grass species (large crabgrass and giant foxtail). Results indicate that among summer annuals, broadleaf species retain larger proportions of their seed compared to grass species at the first opportunity for soybean harvest. As harvest was delayed, more seeds shattered from all species evaluated, indicating timely harvest is critical to maximizing HWSC effectiveness. Studies were conducted on grower fields in Virginia to evaluate the effectiveness of HWSC (field residue and weed seed removal). Results indicate that HWSC can significantly reduce populations of Italian ryegrass in wheat and common ragweed in soybean in the next growing season, but reductions were not observed for Palmer amaranth in soybean. Investigating IWM system for common ragweed control in soybean, HWSC was found to be less effective than soybean planting date (i.e. double cropping after wheat) at reducing common ragweed populations. However, the effectiveness of HWSC varied by location. If HWSC adoption were to become widespread, weeds could adapt by shedding seed earlier in the season. Research was conducted by growing Palmer amaranth populations from across the eastern US in a common garden. Currently there are differences in flowering time and seed shatter among Palmer amaranth populations based on the location of the maternal population, indicating potential for adaptation. This research demonstrates that HWSC is a viable option for weed management in US cropping systems but needs to be stewarded like any other weed management tool. Doctor of Philosophy Herbicide resistance in weeds is a growing problem in the US and around the world. Alternative methods of weed control must be adopted to maintain crop yields in the presence of herbicide-resistant weeds. Researchers and extension specialists strongly advise growers to adopt an integrated weed management (IWM) approach. Integrated weed management involves implementing multiple weed control tactics during a growing season. By using multiple methods of weed control within a given season the chances of weeds becoming resistant or adapting to any single tactic is reduced. Harvest weed seed control (HWSC) is a new tactic developed in Australia in response to herbicide resistance. HWSC targets weed seeds retained on the plant at crop harvest. In a normal crop harvest, the combine removes the grain and spreads crop residues (leaves, stalks, and other plant parts), including weed seeds, back across the field. When HWSC is implemented, weed seeds are destroyed (narrow windrow burning, cage mills) or concentrated and potentially removed from the field (chaff carts, direct bale, chaff lining). Thus, HWSC limits the number of weed seeds returned to the soil seed bank. There is limited research on HWSC and its integration with other tactics, in US cropping systems. For HWSC to be effective it is necessary for weed seeds to be retained on the mother plant in sufficient quantities at crop harvest. Research was conducted in Virginia to determine when weed seeds are shattered during the soybean growing season for 6 economically important weed species, four broadleaf (redroot pigweed, common ragweed, common lambsquarters, and common cocklebur) and two grass species (large crabgrass and giant foxtail). The broadleaf species retained >85% of their seed until the first opportunity for soybean harvest (mid-October). In the grass species, more seed shattered prior to soybean harvest with 50% of large crabgrass and 74% of giant foxtail seed being retained at the first opportunity for soybean harvest. When harvest was delayed seed continued to shatter and less was captured using HWSC. This research indicates broadleaf species are more suitable candidates for HWSC than grass species, among summer annuals. Further research on the ability of seed to germinate in relation to when seeds were shed was conducted on redroot pigweed, common ragweed and common lambsquarters. Results indicate that there are variable effects on germination of these species depending on when they were shed. HWSC was implemented on grower fields to assess the impact on weed populations of 3 weed species (Italian ryegrass, common ragweed, and Palmer amaranth). These experiments compared conventional harvest and HWSC (field residue and weed seed removal) when all other management strategies were the same within that field. Italian ryegrass tiller density in wheat varied by location but was reduced up to 69% in the spring following implementation of HWSC. By wheat harvest, HWSC reduced Italian ryegrass seed head density 67% at one location compared to conventional harvest. In soybean, common ragweed densities were reduced by 22 and 26% prior to field preparation and postemergence herbicide applications, respectively, in the HWSC plots compared to the conventional harvest plots. No differences were observed in common ragweed density by soybean harvest. No differences were observed with Palmer amaranth densities at any point during the soybean growing season. This research show that HWSC can reduce weed populations but is variable and additional research is still needed. IWM experiments were established across Virginia to compare soybean planting date (full season or double cropped), + cover crop (cereal rye/wheat or no cover), and + HWSC (field residue removal) to evaluate the best management strategy for common ragweed in soybean. Across all locations, double cropping soybean behind wheat had the greatest impact on common ragweed densities at the end of the first season. The impact of double cropping soybeans on common ragweed population is due to the emergence pattern of common ragweed; majority of common ragweed emerges prior to planting double cropped soybean (mid-June to early-July). HWSC was variable and only reduced common ragweed density at one of three locations. Widespread adoption of HWSC could place a selection pressure on weeds to shatter seed earlier in the season. A common garden experiment was conducted in Blacksburg, VA to assess Palmer amaranth populations collected from central Florida to southern Pennsylvania for differences in flowering time, time to seed shatter, and other phenotypic traits. Results indicate that latitude of the maternal population influences time to first flower with a 0.53 d reduction in flowering time for every degree north in latitude the maternal population was collected from. The strongest predictor of Palmer amaranth flowering time was emergence date/daylength. For every day emergence was delayed the time to first flower was reduced by 0.31 and 0.24 d for female and male plants, respectively. Time from emergence or first flower to first seed shatter was reduced by 0.48 or 0.17 d, respectively, for each day emergence was delayed. These results indicate that differences exist currently among Palmer amaranth populations and the selection pressure of HWSC could push these populations to flower and shatter seed early.

Even under the best management strategies contemporary crops face yield losses from diverse threats such as, pathogens, pests, and environmental stress. Adding to this management challenge is that under current global climate projections these impacts are predicted to become even greater. Natural genetic variation, long used by traditional plant breeders, holds great promise for adapting high performing agronomic lines to these stressors. Yet, efforts to bolster crop plant resilience using wild relatives have been hindered by time consuming efforts to develop genomic tools and/or identify the genetic basis for agronomic traits. Thus, increasing crop plant resilience requires developing and deploying approaches that leverage current high-throughput sequencing technologies to more rapidly and robustly develop genomic tools in these systems. Here we report the integration of bioinformatic and statistical tools to leverage high-throughput sequencing to 1) develop a machine learning approach to determine factors impacting transcriptome assembly and quantitatively evaluate transcriptome completeness, 2) dissect complex physiological pathway interactions in Solanum pimpinellifolium under combined stresses—using comparative transcriptomics, and 3) develop a genome assembly pipeline that can be deployed to rapidly assemble a more contiguous genome, unraveling previously hidden complexity, using Phytopthora capsici as a model. As a result, we have generated strategic guidelines for transcriptome assembly and developed an orthologue and reference free, machine learning based tool "WWMT" to quantitatively score transcriptome completeness from short read data. Secondly, we identified "hub genes" and describe genes involved with "cross-talk" between drought and herbivore stress response pathways. Finally, we demonstrate a protocol for combining long-read sequencing from the Oxford Nanopore Technologies MinION, and short-read data, to rapidly assembly a cost-effective, contiguous and relatively complete genome. Here we uncovered hidden variation in a well-known plant pathogen finding that the genome was 92% bigger than previous estimates with more than 39% of duplicated regions, supporting a hypothesized recent whole genome duplication in this clade. This community resource will support new functional and evolutionary studies in this economically important pathogen. Doctor of Philosophy Meeting the food production demands of a burgeoning population in a changing environment, means adapting crop plants to become more resilient to environmental stress. One of the greatest barriers to understanding and predicting crop responses to future environmental change is our poor understanding of the functional and genomic basis of stress resistance traits for contemporary crops. This impediment presents a barrier for rapid crop improvement technologies, such as, gene editing or genomic selection, that is only partially overcome by generating large amounts of sequencing data. Here we need tools that allow us to process and evaluate huge amounts of data generated from next generation sequencing studies to help identify genomic regions associated with agronomic traits. We also need technical approaches that allow us to disentangle the complex genetic interactions that drive plant stress responses. Here we present work that used statistical analysis and recent advances of artificial intelligence to develop a bioinformatic approach to evaluate genomic sequencing data prior to downstream analyses. Secondly, we used a reductionist approach to filter thousands of genes to key genes associated with combined stress responses (herbivory and drought), in the most widely used vegetable in the world, tomato. Finally, we developed a method for generating whole genome sequences that is low-cost and time sensitive and tested it using a well-known plant pathogen genome, wherein we unraveled significant hidden complexity. Overall this work provides community-wide genomic tools and information to promote crop resilience.

Soybean is an important source of protein in animal feed, and growing demand for meat consumption worldwide has led to increased soybean production. Over 120 million metric tons of soybean were harvested in the United States in 2018, approximately one-third of the world production. In the Mid-Atlantic region, soybean is one of the most valuable field crops. Major foliar diseases that reduce soybean yield in the Mid-Atlantic region are frogeye leaf spot (FLS) and Cercospora leaf blight. In addition to crop rotation and host resistance, foliar fungicides, often with quinone outside inhibitor (QoI) active ingredients, are used to manage these soybean foliar diseases. Yield benefits of foliar fungicides have been inconsistent and this may be the result of low disease pressure, unfavorable environmental conditions for disease development, or the presence of fungal pathogen populations that have developed resistance to fungicides. The objectives of this research were 1) to develop a pyrosequencing-based assay to rapidly quantify QoI resistance frequencies in Cercospora sojina, the causal agent of FLS, 2) to examine the effects of fungicide application timings, disease pressure, and environmental factors on soybean yield, and 3) to develop a weather-based soybean foliar fungicide application decision aid for the Mid-Atlantic U.S. using a threshold decision rule. A pyrosequencing assay targeting the G143A mutation was designed, and a Virginia survey of C. sojina populations indicated that the G143A mutation conferring QoI resistance is widespread. In small plot fungicide application timing experiments, five weekly fungicide applications starting at beginning pod (R3) resulted in the greatest yield, but for single fungicide applications, R3 or 1 week after R3 resulted in the greatest yields. There was positive relationship between the cumulative number of disease favorable days (mean daily temperature 20-30°C and ≥ 10 hours of relative humidity >90%) from planting to R3 and disease severity at the full pod stage (r = 0.97, P =

Hyaloperonospora arabidopsidis (Hpa) is a naturally occurring oomycete pathogen on Arabidopsis thaliana. It is related to downy mildews of economically important crops such as cabbage, kale and broccoli, belonging to the Brassicaceae family. Downy mildew pathogens are obligate biotrophs that extract nutrients exclusively from living plant cells. As a part of its obligate biotrophy lifestyle, Hpa has lost the ability to assimilate inorganic nitrogen and sulfur. It thus has to acquire these nutrients from the host in an organic form; possibly amino acids. Using a reverse genetic approach, I was able to identify two host amino acid transporters that are up-regulated during Hpa infection: AAP3 and AAP6. Both of these transporters are localized in the vasculature of the plant, AAP3 mostly in the root, and AAP6 in the roots and shoots. Using transgenic lines of Arabidopsis containing transcriptional and translational reporter fusion constructs for these genes, I found that AAP3 displays increased mRNA accumulation which is attributable to an increased promoter activity in regions of shoot tissue colonized by Hpa. On the other hand, AAP6 displays a mild increase in mRNA accumulation under Hpa infection, but the induction becomes more prominent at the protein level as seen by fluorescence from GFP fused to AAP6. Surprisingly, null mutants of AAP3 did not impact Hpa growth whereas null mutants of AAP6 made the plant more susceptible to Hpa. Furthermore, aap6 mutants accumulate fewer free amino acids in the phloem compared to wild-type plants when infected with Hpa. Together, these results suggest that AAP6 acts a nutritional starvation gene for the pathogen and hence aids the plant during infection. While we now know more about AAP3's regulation during infection, its function remains to be elucidated. To successfully colonize a plant, a pathogen must be able to achieve both suppression of plant immunity and acquisition of nutrients from the plant host. While the former has been well studied, research on the latter is sparse. This work was a step in the direction to increase our understanding of potential players in nutrient acquisition by pathogens. Doctor of Philosophy A key aspect of achieving and maintaining food security is sustainable agricultural production. This is endangered by plant diseases that lead to large losses in crop production. All plant pathogens have to acquire food and nutrients from the plants they infect. Understanding how they acquire nutrients from the plant at a molecular level can give us insight into potential methods to prevent this and hence reduce the impact of plant diseases. One such nutrient is nitrogen. Nitrogen is essential to all of an organism’s cellular and metabolic processes. Organisms utilize nitrogen by converting it from inorganic forms such as nitrates to organic forms such as amino acids. Some plant pathogens, such as Hyaloperonospora arabidopsidis (Hpa), which causes downy mildew disease on the model plant Arabidopsis, complete their entire life cycle on a living plant. They are also unable to convert the inorganic nitrogen to organic forms and hence depend on acquiring organic forms of nitrogen from the plant. Thus, it is important to understand how they acquire amino acids from the plant. Plants use amino acid transporters that serve as a siphon or a pump in moving amino acids from one region of the plant to another. It is possible that pathogens manipulate plant’s amino acid transporters to move amino acids towards the infection site while, at the same time, plants might use another set of transporters to move amino acids away from the pathogen. This work was an attempt at understanding this potential role of plant amino acid transporters in plant-pathogen interactions using the model system of Hpa and Arabidopsis.

A growing body of circumstantial evidence suggests that ice nucleation active (Ice(+)) bacteria contribute to the initiation of precipitation by heterologous freezing of super-cooled water in clouds. However, little is known about the concentration of Ice(+) bacteria in precipitation, their genetic and phenotypic diversity, and their relationship to air mass trajectories and precipitation chemistry. In this study, 23 precipitation events were collected over 15 months in Virginia, USA. Air mass trajectories and water chemistry were determined and 33 134 isolates were screened for ice nucleation activity (INA) at -8 °C. Of 1144 isolates that tested positive during initial screening, 593 had confirmed INA at -8 °C in repeated tests. Concentrations of Ice(+) strains in precipitation were found to range from 0 to 13 219 colony forming units per liter, with a mean of 384±147. Most Ice(+) bacteria were identified as members of known and unknown Ice(+) species in the Pseudomonadaceae, Enterobacteriaceae and Xanthomonadaceae families, which nucleate ice employing the well-characterized membrane-bound INA protein. Two Ice(+) strains, however, were identified as Lysinibacillus, a Gram-positive genus not previously known to include Ice(+) bacteria. INA of the Lysinibacillus strains is due to a nanometer-sized molecule that is heat resistant, lysozyme and proteinase resistant, and secreted. Ice(+) bacteria and the INA mechanisms they employ are thus more diverse than expected. We discuss to what extent the concentration of culturable Ice(+) bacteria in precipitation and the identification of a new heat-resistant biological INA mechanism support a role for Ice(+) bacteria in the initiation of precipitation.

Poison ivy (Toxicodendron radicans (L.) Kuntze) is a native perennial liana widely recognized for the production of urushiol, and the associated contact dermatitis it causes in humans. Poison ivy is predicted to become both more prevalent and more noxious in response to projected patterns of global change. Moreover, poison ivy is an important food source for avian species, and urushiol has numerous applications as a high-value engineering material. Thus, this curious weed has many avenues for future concern, and promise. Here, I address gaps in knowledge about poison ivy ecology and physiology so that we may better understand its weediness and utilize its benefits. I address three core areas: poison ivy establishment patterns; biotic interactions with multiple taxa; and the development of molecular tools for use in poison ivy. I found that the early life stage of seedling emergence is a critical linchpin in poison ivy establishment due largely to herbivore pressure from large grazers. I also describe the multifaceted relationship between poison ivy and avian frugivores that not only disperse the drupes of poison ivy but also aid in reduction of fungal endophytic phytopathogens. A survey of poison ivy urushiols yielded that while variation in urushiol congeners was high across individuals, relative congener levels were stable within individuals over a two month period. Lastly I demonstrate best practices for introducing and transiently expressing recombinant DNA in poison ivy as a step towards future reverse genetic procedures. Doctor of Philosophy Poison ivy is a native plant best known for its capacity to cause allergenic skin reactions in humans due to the chemical urushiol, which is found in all parts of the plant. While most people prefer to avoid this plant, poison ivy is an important food source for birds. In addition, urushiol has numerous applications as an engineered material. Despite these positive aspects, poison ivy is among those plants that are responding well to global change, such as increasing CO₂ levels and habitat fragmentation. Poison ivy has been shown to increase in size and produce more allergenic forms of urushiol under elevated CO₂ levels and there are concerns that poison ivy prefers the disturbed areas created by habitat fragmentation. These attributes suggest that poison ivy will become more prevalent and more noxious in the coming years. Thus, this curious weed has many avenues for both future concern and promise. To aid in our ability to manage poison ivy in the future, I used a combination of field, greenhouse, and laboratory studies to study the ecology of poison ivy. I investigated the early stages of the poison ivy life cycle, and the relationship between poison ivy and the animals that interact with it. I found that the earliest life stages of poison ivy are a critical linchpin for poison ivy survival due largely to large animals like deer eating the seedlings. I also describe the multifaceted relationship between poison ivy and birds, which not only disperse the seeds of poison ivy but also aid in reducing pathogens associated with the seeds. I surveyed the amounts and types of urushiols that poison ivy produces and found them to be highly variable from plant to plant, but relatively stable over time within a plant. Lastly, I demonstrate best practices for transient transgene expression in poison ivy leaves as a step towards future genetic studies. These studies help expand our understanding of a problematic weed, and pave the way for future studies in weed ecology and in the utilization of urushiol in positive applications, showing that even poison ivy can be of benefit to the environment and humans.

Industry demand for local sources of grain for animal feed has increased sorghum production in the mid-Atlantic region of the U.S. Sorghum anthracnose (causal agent Colletotrichum sublineola) and the grain mold complex, which includes mycotoxin-producing Fusarium spp., limit the yield and quality of grain sorghum in humid climates worldwide. A majority of U.S. grain sorghum production is in arid regions, and management strategies have not been developed for the mid-Atlantic U.S. where warm, wet conditions favor disease. The specific objectives of this research were to: (1) determine the effectiveness of fungicides and their application timing for the management of sorghum foliar anthracnose, (2) compare five grain sorghum hybrids for their susceptibility to foliar anthracnose, grain mold and mycotoxin contamination under field conditions, (3) integrate host resistance and fungicide application to manage anthracnose and grain mold, and (4) identify Fusarium spp. associated with grain mold and mycotoxin contamination of sorghum in the mid-Atlantic U.S. For Objective 1, it was determined that a single application of pyraclostrobin-containing fungicide no later than flowering reduced anthrancose, protected yield and maximized farm income. Objective 2 focused on sorghum hybrid selection as a disease management tactic, and it was determined that hybrids with high yield potential and moderate disease resistance should be selected for mid-Atlantic sorghum production in order to maximize grain yield and quality while minimizing the need for fungicide inputs. Objective 3 focused on integrated management and demonstrated that under moderate disease pressure, a high-yielding susceptible hybrid required a single application of pyraclostrobin-based fungicide to minimize fungal diseases and maintain acceptable yields, whereas under high disease pressure it was necessary to integrate hybrid resistance and judicous applications of fungicides. The aim of Objective 4 was to characterize potential causal agents of mycotoxin contamination in mid-Atlantic sorghum, and thirteen phylogenetically distinct Fusarium species (F. lacertarum, F. graminearum. F. armeniacum, F. proliferatum, F. fujikuroi, F. verticillioides, F. thapsinum and several in Fusarium incarnatum-equiseti species complex) were found to be associated with grain mold and fumonisin and/or deoxynivalenol contamination of sorghum grain. This work has provided insights into the impacts of fungal diseases on grain sorghum yield and quality in the mid-Atlantic and has aided in development of best management practices for the region. Doctor of Philosophy Sorghum is grown in tropics, sub-tropics and semi-arid region worldwide for food, feed, forage and fuel. Sorghum acreage in the mid-Atlantic is increasing due the demand for locally grown grain by poultry and swine industries. During the growing season, warm and humid conditions are common in the southeastern and mid-Atlantic states favoring fungal diseases development that reduce the grain yield and quality. Anthracnose and grain mold, which includes toxic mycotoxin-producing Fusarium species, are the two major constraints in sorghum production in the region. However, management alternatives have not been developed. The main goal of this research was to develop management strategies to protect yield and maximize farm profitability by controlling anthracnose and grain mold of sorghum using chemicals and/or host resistance. The specific objectives were to: (1) determine the effectiveness of fungicides and their application timing for the management of sorghum foliar anthracnose, (2) compare grain sorghum hybrids for their susceptibility to foliar anthracnose, grain mold and mycotoxin contamination under field conditions, (3) assess the value of integrating host resistance and judicious use of fungicides to manage sorghum anthracnose and grain mold, and (4) identify Fusarium spp. associated with grain mold and mycotoxin contamination of sorghum in the mid-Atlantic U.S. Results from this research indicate that a single application of pyraclostrobin-containing fungicides no later than flowering reduces anthrancose, protects yield, and increases farm income. Sorghum hybrids varied in susceptibility to anthrancnose and grain mold, and planting a moderately resistant hybrid and applying a fungicide under high disease risk conditions provided the greatest return on investment. Both fumonisin and deoxynivalenol were frequently detected from sorghum grain, and mycotoxin contamination was associated with 13 different Fusarium species from three distinct species complexes. Based on the results of this work, best management practices for minimizing sorghum disease losses were developed for the mid-Atlantic region.

Colletotrichum acutatum and C. gloeosporioides are fungal plant pathogens that have a global distribution, extensive host range, and convoluted taxonomy. Both species can cause grape ripe rot and are considered endemic to Virginia US. In 2012, C. acutatum and C. gloeosporioides were reclassified into species complexes that consist of 31 and 22 accepted species, respectively. The objectives of this study were to: 1) survey Virginia vineyards for grape ripe rot, and morphologically and phylogenetically identify isolates to the species within the complexes, 2) conduct an in vitro fungicide assay to screen fifteen commercial fungicides and combinations of two fungicides for efficacy to control isolates from seven Colletotrichum species from Virginia vineyards, 3) sequence gene fragments from three subunits of the SDH enzyme in the fungicide-screened isolates to observe potential resistance mutations, 4) investigate the susceptibility of three grapevine tissues to Colletotrichum species, 5) observe potential infection structures before and after the application of fungicides, 6) evaluate the efficacy of commercial fungicide controls of grape ripe rot in the field, and determine the most advantageous timing of applications. In my studies, I identified six Colletotrichum species: C. aenigma, C. conoides, C. fioriniae, C. gloeosporioides, C. kahawae, and C. nymphaeae. I also found two additional groups; an isolate similar to C. limetticola and C. melonis and a group of isolates that are similar to C. alienum, C. fructicola, and C. nupharicola. I also identified captan, and mancozeb as two potential active ingredients for control of grape ripe rot isolates from Virginia via the in vitro fungicide assay. Additionally, I found that combinations of two active ingredients could increase the efficacy of benzovindiflupyr, copper, and polyoxin-D. C. fioriniae germination and production of melanized appressoria was documented on leaves. I observed appressorium formation with isolates of two C. fructicola-like genotypes and C. nymphaeae, as well as secondary conidiation with isolates of C. aenigma, C. fructicola-like genotype 3, and C. nymphaeae on blooms. And finally, benzovindiflupyr, cyprodinil + fludioxonil pre-mix, and potassium phosphite + tebuconazole were identified as candidates for chemical control for grape ripe rot in the field. PHD Colletotrichum acutatum and C. gloeosporioides are two fungal plant pathogens that are found on a wide range of crops around the globe. Both fungal species cause the disease grape ripe rot and have been found in Virginia (VA) USA since the late 1800s. Originally, grape ripe rot was considered a minor disease in VA; however, based on communications with local VA vineyard managers, grape ripe rot was found to cause up to 30% direct crop loss. Further indirect economic loss occurs during wine production due to the production of unpalatable, tobacco-like, off flavors from the infected grapes. Sensory studies found this wine flavor change occurred with as little as 3% of the total crushed grapes being infected. Grape ripe rot appears as a sunburn-like, tan injury on the surface of white-fruited grape berries. As the disease progresses, the dark injury expands across the surface of the berry and rings of salmon-colored spore masses form. On red-fruited grapes, the formation of spore masses is usually the first observable sign. Over time, the infected berries will shrivel down to a soft, pustule-covered raisin. Both C. acutatum and C. gloeosporioides cause the same grape ripe rot symptoms on fruit and overlap in fungal appearance. In addition, investigations of these pathogens using molecular techniques have revealed that each consists of a number of genetically distinct groups that are difficult to distinguish by appearance. Therefore, in 2012, C. acutatum and C. gloeosporioides were reclassified into 31 and 22 newly accepted species, respectively, using molecular techniques. The objectives of this study were to: 1) survey VA vineyards for grape ripe rot, and visually and molecularly identify isolates to the species within the new complexes, 2) conduct a laboratory fungicide assay to screen fifteen commercial fungicides and combinations of two fungicides for control of isolates from VA vineyards, 3) sequence gene fragments from three subunits of the SDH enzyme in the fungicide-screened isolates to observe potential resistance mutations, 4) investigate the susceptibility of three grapevine tissues to Colletotrichum species, 5) observe potential infection structures before and after the application of fungicides, 6) evaluate the efficacy of commercial fungicide controls of grape ripe rot in the field, and determine the most advantageous timing of applications. In my studies, I identified six Colletotrichum species: C. aenigma, C. conoides, C. fioriniae, C. gloeosporioides¸ C. kahawae, and C. nymphaeae. I also found two additional groups; an isolate similar to C. limetticola and C. melonis and a group of isolates that are similar to C. alienum, C. fructicola, and C. nupharicola. Our lab also identified four active ingredients as potential controls of grape ripe rot in the laboratory fungicide assay; captan, mancozeb, tetraconazole and thiophanate-methyl. Additionally, combinations of two compounds can increase the effectiveness of benzovindiflupyr, copper, and polyoxin-D. of C. fructicola-like isolates, and C. nymphaeae formed infection structures on blooms. of C. aenigma, C. fructicola-like genotype 3, and C. nymphaeae formed spores on blooms without producing symptoms. C. fioriniae spores germinated and produced infection structures on leaves without producing symptoms.

Palmer amaranth (Amaranthus palmeri) and common ragweed (Ambrosia artemisiifolia) can cause detrimental soybean yield loss. Due to widespread resistance to glyphosate and ALS-inhibiting herbicides, growers rely on protoporphyrinogen oxidase inhibiting herbicides (PPO) such as flumioxazin applied preemergence (PRE) and fomesafen postemergence (POST) to control both weeds. Experiments were conducted with the overarching goal of reducing PPO selection pressure for Palmer amaranth and common ragweed. Flumioxazin alone PRE controlled Palmer amaranth near 100%. However, sulfentrazone combined with pyroxasulfone or pendimethalin provided similar control to flumioxazin. Acetochlor and linuron controlled common ragweed 96%. Glufosinate applied POST controlled Palmer amaranth and common ragweed 74-100%, regardless of PRE treatment. Flumioxazin PRE followed by fomesafen POST controlled common ragweed well; however, several non-PPO herbicide treatments or programs with only 1 PPO-inhibiting herbicide provided similar common ragweed control as the 2 PPO system (flumioxazin followed by fomesafen). Treatments consisting of a PRE and POST herbicide controlled Palmer amaranth at least 80% and common ragweed 95%. To reduce PPO selection pressure, soybean producers growing glufosinate-resistant soybean may use flumioxazin PRE followed by glufosinate POST whereas non-glufosinate-resistant soybean growers should reduce PPO herbicide use by using a non-PPO herbicide PRE. Alternatively, these producers can effectively reduce PPO selection pressure by implementing residual combinations of a PPO-inhibiting herbicide + non-PPO with spectrums of weed control that overlap at either Palmer amaranth or common ragweed. Master of Science in Life Sciences Soybean producers planted 35.4 million hectares in the US during 2018. Palmer amaranth (Amaranthus palmeri) and common ragweed (Ambrosia artemisiifolia) are both common and problematic in soybean production. The introduction of a glyphosate-resistant soybean cultivars coupled with glyphosate allowed soybean producers to easily control these weeds along with many other broadleaf and grass weeds. However, over reliance on glyphosate selected for biotypes of common ragweed and Palmer amaranth resistant to the herbicide. In response, soybean producers have reverted to preemergence (PRE) herbicides and alternative modes of action postemergence (POST) to control these herbicide-resistant weeds. One such herbicide mode of action is inhibition of protoporphyrinogen oxidase (PPO). Flumioxazin and fomesafen are both PPO-inhibiting herbicides and have been widely used in soybean, however increasing reliance on PPOs has selected for resistant common ragweed and Palmer amaranth biotypes. This research focused on reducing risk of PPO-inhibiting herbicide resistance development (“selection pressure”) by finding alternatives to or combinations with PPOinhibiting herbicides that would effectively control both weeds and thus preserve effectiveness of a valuable herbicide group. Of PRE herbicides applied alone, flumioxazin was the only treatment to control Palmer amaranth >79% 14 DA-PRE at Painter 2017. However, combination of PRE herbicides such as sulfentrazone or metribuzin in combination with pyroxasulfone, and pendimethalin + sulfentrazone, all controlled Palmer amaranth well. While metribuzin and pendimethalin alone did not provide as much control, a POST application of glufosinate coupled with these residual herbicides adequately controlled Palmer amaranth. Soybean producers can effectively control Palmer amaranth with a non-PPO PRE herbicide followed by glufosinate postemergence (POST) or residual combinations of a PPO + non-PPO while reducing risk of herbicide resistance development. Several PRE herbicide treatments adequately controlled common ragweed. During 2017, residual herbicides that controlled common ragweed at least 90% included flumioxazin, flumioxazin + clomazone, linuron, or metribuzin, fomesafen + linuron, and linuron + clomazone. All treatments controlled common ragweed greater than 94% during 2018, except metribuzin, linuron, and clomazone, which controlled the weed 75, 86, and 90%, respectively. Fomesafen alone or in combination with metribuzin controlled common ragweed 80 to 84%. Regardless of PRE, glufosinate POST controlled common ragweed 99% 56 and 70 days after planting (DAP). In fields infested with common ragweed yet to develop PPO resistance, growers should use a non-PPO herbicide in combination with flumioxazin PRE. Additionally, tank mixtures of effective MOAs PRE followed by glufosinate rather than a PPO POST may reduce herbicide selection pressure. The final study set out to determine which was more critical to controlling herbicideresistant Palmer amaranth and common ragweed in soybean, a PPO-inhibiting herbicide applied PRE or POST. Flumioxazin applied PRE controlled both weeds almost completely. Acetochlor and linuron did not control common ragweed as well, but controlled Palmer amaranth >96%. Both metribuzin and clomazone were weaker on common ragweed and Palmer amaranth. However, all PRE herbicide treatments followed by glufosinate or fomesafen controlled Palmer amaranth and common ragweed at least 80 and 95%, respectively. To reduce PPO selection pressure, soybean producers growing glufosinate-resistant soybean may use flumioxazin PRE followed by glufosinate POST whereas non-glufosinate-resistant growers should reduce PPO herbicide use by using a non-PPO herbicide PRE. Alternatively, these producers can effectively reduce PPO selection pressure by implementing residual combinations of a PPO-inhibiting herbicide + non-PPO with spectrums of weed control that overlap at either Palmer amaranth or common ragweed. Results from these experiments suggest PPO-inhibiting herbicides are critical for common ragweed and Palmer amaranth control. Previous research has shown effective tank mixtures with various effective MOAs has reduced the risk of herbicide resistance development. Protoporphyrinogen oxidase herbicides should be used sparingly and in combination with effective non-PPO herbicides to reduce selection pressure. Soybean producers planted 35.4 million hectares in the US during 2018. Palmer amaranth (Amaranthus palmeri) and common ragweed (Ambrosia artemisiifolia) are both common and problematic in soybean production. The introduction of a glyphosate-resistant soybean cultivars coupled with glyphosate allowed soybean producers to easily control these weeds along with many other broadleaf and grass weeds. However, over reliance on glyphosate selected for biotypes of common ragweed and Palmer amaranth resistant to the herbicide. In response, soybean producers have reverted to preemergence (PRE) herbicides and alternative modes of action postemergence (POST) to control these herbicide-resistant weeds. One such herbicide mode of action is inhibition of protoporphyrinogen oxidase (PPO). Flumioxazin and fomesafen are both PPO-inhibiting herbicides and have been widely used in soybean, however increasing reliance on PPOs has selected for resistant common ragweed and Palmer amaranth biotypes. This research focused on reducing risk of PPO-inhibiting herbicide resistance development (“selection pressure”) by finding alternatives to or combinations with PPOinhibiting herbicides that would effectively control both weeds and thus preserve effectiveness of a valuable herbicide group. Of PRE herbicides applied alone, flumioxazin was the only treatment to control Palmer amaranth >79% 14 DA-PRE at Painter 2017. However, combination of PRE herbicides such as sulfentrazone or metribuzin in combination with pyroxasulfone, and pendimethalin + sulfentrazone, all controlled Palmer amaranth well. While metribuzin and pendimethalin alone did not provide as much control, a POST application of glufosinate coupled with these residual herbicides adequately controlled Palmer amaranth. Soybean producers can effectively control Palmer amaranth with a non-PPO PRE herbicide followed by glufosinate postemergence (POST) or residual combinations of a PPO + non-PPO while reducing risk of herbicide resistance development. Several PRE herbicide treatments adequately controlled common ragweed. During 2017, residual herbicides that controlled common ragweed at least 90% included flumioxazin, flumioxazin + clomazone, linuron, or metribuzin, fomesafen + linuron, and linuron + clomazone. All treatments controlled common ragweed greater than 94% during 2018, except metribuzin, linuron, and clomazone, which controlled the weed 75, 86, and 90%, respectively. Fomesafen alone or in combination with metribuzin controlled common ragweed 80 to 84%. Regardless of PRE, glufosinate POST controlled common ragweed 99% 56 and 70 days after planting (DAP). In fields infested with common ragweed yet to develop PPO resistance, growers should use a non-PPO herbicide in combination with flumioxazin PRE. Additionally, tank mixtures of effective MOAs PRE followed by glufosinate rather than a PPO POST may reduce herbicide selection pressure. The final study set out to determine which was more critical to controlling herbicideresistant Palmer amaranth and common ragweed in soybean, a PPO-inhibiting herbicide applied PRE or POST. Flumioxazin applied PRE controlled both weeds almost completely. Acetochlor and linuron did not control common ragweed as well, but controlled Palmer amaranth >96%. Both metribuzin and clomazone were weaker on common ragweed and Palmer amaranth. However, all PRE herbicide treatments followed by glufosinate or fomesafen controlled Palmer amaranth and common ragweed at least 80 and 95%, respectively. To reduce PPO selection pressure, soybean producers growing glufosinate-resistant soybean may use flumioxazin PRE followed by glufosinate POST whereas non-glufosinate-resistant growers should reduce PPO herbicide use by using a non-PPO herbicide PRE. Alternatively, these producers can effectively reduce PPO selection pressure by implementing residual combinations of a PPO-inhibiting herbicide + non-PPO with spectrums of weed control that overlap at either Palmer amaranth or common ragweed. Results from these experiments suggest PPO-inhibiting herbicides are critical for common ragweed and Palmer amaranth control. Previous research has shown effective tank mixtures with various effective MOAs has reduced the risk of herbicide resistance development. Protoporphyrinogen oxidase herbicides should be used sparingly and in combination with effective non-PPO herbicides to reduce selection pressure.

Cover crops have become an important part of cropping systems in the United States, especially in the Mid-Atlantic region. Rapeseed is a popular choice due to its deep growing taproot which creates soil macropores and increases water infiltration. If not properly terminated rapeseed can become problematic due to its pod-shattering tendency and its difficulty to terminate with herbicides once it enters reproductive growth. Results indicate termination of rapeseed is most effective when the cover crop is small. Combinations that successfully terminated rapeseed include glyphosate plus 2,4-D and paraquat plus 2,4-D. Halauxifen-methyl is a new Group 4 herbicide marketed for preplant burndown horseweed (Conyza canadensis L.) control. Previous research indicates that halauxifen effectively controls glyphosate-resistant horseweed. However, little is known about control of other common winter annual weeds by halauxifen. Results indicate halauxifen has a narrow spectrum of control providing adequate control (>80%) of horseweed, henbit (Lamium amplexicaule L.), and purple deadnettle (Lamium purpureum L.), while failing to control cutleaf evening-primrose (Oenothera laciniata Hill), curly dock (Rumex crispus L.), purple cudweed (Gamochaeta purpurea L. Cabrera), common chickweed (Stellaria media L.), and mousear chickweed (Cerastium L.). Little is known of cotton (Gossypium hirsutum L.) tolerance to halauxifen applied preplant burndown. Results indicate cotton is more tolerant to halauxifen than 2,4-D or dicamba when the interval between preplant application and cotton planting is less than 30 days. Master of Science in Life Sciences Cover crops are an important part of cropping systems in the United States, especially in the Mid-Atlantic region. Producers utilize cover crops to aid in weed suppression, reduce soil erosion, as well as to increase soil health. Cereals, legumes, and Brassicaceae species are popular cover crops planted either as monocultures or mixtures. Rapeseed can become problematic due to its difficulty to terminate once it enters reproductive stage, as well as its podshattering characteristic. Experiments were conducted to evaluate various herbicides and herbicide combinations for rapeseed termination two application timings. At three locations where rapeseed averaged 12 cm in height at early termination, and 52 cm in height at late termination, glyphosate + 2,4-D was most effective, controlling rapeseed (96%) 28 days after early termination (DAET). Paraquat + atrazine + atrazine (92%), glyphosate + saflufenacil (91%), glyphosate + dicamba (91%), and glyphosate (86%) all provided at least 80% control 28 DAET. Paraquat + 2,4-D (85%), glyphosate + 2,4-D (82%), and paraquat + atrazine + mesotrione (81%) were the only treatments to provide at least 80% control 28 days after late termination (DALT). At one location where rapeseed was much taller (41 cm early termination; 107 cm late termination), herbicides were much less effective, as no herbicide treatments provided greater than 80% control. Results indicated that rapeseed size at time of termination was more critical to successful termination than herbicide choice. Prior to the development of glyphosate-resistant horseweed, producers were able to control horseweed and other weeds with glyphosate applied preplant burndown. Producers now rely on auxin herbicides tank mixed with glyphosate and a residual herbicide to control horseweed and other winter weeds prior to cash crop planting. Experiments were conducted to evaluate halauxifen-methyl, a new Group 4 herbicide, for control of horseweed and other commonly encountered winter annual weeds. Halauxifen (89%) controlled small horseweed (15 cm in height at time of application) than either dicamba (77%) or 2,4-D evaluated (64%). Halauxifen provided adequate control (>80%) of henbit (Lamium amplexicaule L). and purple deadnettle (Lamium purpureum L.), while failing to effectively control of cutleaf evening-primrose (Oenothera laciniata Hill), curly dock (Rumex crispus L.), purple cudweed (Gamochaeta purpurea L. Cabrera), common chickweed (Stellaria media L. Vill.), and mousear chickweed (Cerastium L.). Results indicate that halauxifen has a narrow spectrum of control and should be tank mixed with 2,4-D or glyphosate in order to control weeds other than horseweed and henbit. Glyphosate plus dicamba or 2,4-D plus a residual herbicide is typically applied prior to cotton planting. Previous research has shown that as long as rainfall requirements and rotation intervals are met, no adverse effects on cotton is observed from 2,4-D or dicamba herbicides. Little is known of cotton tolerance to halauxifen applied preplant burndown. Experiments were conducted to determine if halauxifen applied sooner than the labeled 30-day rotation interval would injure cotton. Very little injury was observed from halauxifen (9%) applied at-planting, however dicamba (26%) and 2,4-D (21%) applied at the same timing did injure cotton. Auxin herbicides applied earlier in the season resulted in little injury (

Weed control and desiccation are routinely executed with herbicides. Potato vine desiccation facilitates harvest, improves skin set, and regulates tuber size. Saflufenacil, glufosinate, saflufenacil plus glufosinate, and carfentrazone plus glufosinate were compared to diquat applied at 43, 31, and 17% B potatoes; similar vine desiccation (14 days after treatment), skin set, and yield were noted amongst treatments. Residual herbicides are routinely used for weed control in pumpkin. Fluridone and acetochlor formulations applied preemergence were evaluated in direct-seeded pumpkin compared to other labeled herbicides. Fluridone resulted in total crop loss following heavy rainfall immediately after planting; less rainfall resulted in transient injury. Acetochlor formulations resulted in significant pumpkin injury (34 to 39%) 14 days after planting. S-metolachlor controlled weeds similar to acetochlor without significant injury. Palmer amaranth has developed resistance to six different herbicide modes of action. The weed grows rapidly and is best controlled

Dollar spot is one of the most common diseases of warm- and cool-season turfgrass stands and is especially devastating on creeping bentgrass (Agrostis stolonifera L.). The fungus Sclerotinia homoeocarpa degrades the foliage by creating silver, dollar-sized depressions of dead and bleached turf. Frequent fungicide applications and cultural management strategies are required throughout the growing season to prevent or reduce severity of this disease. Previous research has demonstrated that ferrous sulfate applied at 48.8 kg ha-1 suppresses dollar spot epidemics without traditional fungicides. In vitro studies showed 100 to 1,000 mg kg-1 of ferrous sulfate directly suppressed S. homoeocarpa growth of an isolate collected from an established, intensively-maintained creeping bentgrass putting green. Genetic diversity of S. homoeocarpa segregates isolates into two groups; strains generally associated with warm-season and cool-season grasses. It is unknown whether isolates of each group react similarly in the presence of ferrous sulfate. Our research explored use rates of ferrous sulfate required to suppress 50% of dollar spot in the field and in vitro. Ferrous sulfate (heptahydrate 20% Fe, Valudor Products Inc) rates in field trials included 0, 4.88, 24.4, 48.8, and 97.6 kg ha-1. Our results indicate a hyperbolic relationship between ferrous sulfate rate and dollar spot reduction. Using this model, 26.4 kg ha-1 reduced dollar spot incidence by 50%. We concluded that ferrous sulfate suppresses 50% of S. homoeocarpa mycelial growth at between 480 and 720 mg L-1 concentration in 0.25 strength potato dextrose agar in vitro, and fungitoxic activity of ferrous sulfate was dependent primarily on historical fungicide inputs at isolate collection sites. The use of ferrous sulfate may supplement traditional fungicide use. Chlorothalonil is the most common fungicide used to suppress dollar spot in turfgrass. Annual site-use limitations of chlorothalonil often prevent turf managers from achieving acceptable dollar spot control throughout the season. It is not known how ferrous sulfate may contribute to a successful chlorothalonil fungicide program. Therefore, we examined whether ferrous sulfate can be used to minimize chlorothalonil requirements through reducing active ingredient concentrations and extending the longevity, while still maintaining acceptable disease control. Chlorothalonil treatments were applied at 0, 2.28, 4.57, 6.86, and 9.16 kg ai ha-1 (Daconil WeatherStik) across plots treated with and without 48.8 kg ha-1 ferrous sulfate applied bi-weekly. Ferrous sulfate reduced the chlorothalonil rates necessary for 80% disease reduction by 36 to 51% across all trials. Additional studies showed that ferrous sulfate applied with chlorothalonil increased duration of disease control by five days and eliminated two seasonal treatments. Our research expands the guidelines for practical ferrous sulfate usage for dollar spot suppression by elucidating the rate-to-disease relationship and providing best management practices involving admixtures with chlorothalonil. Master of Science Turfgrass systems offer many environmental and recreational benefits. Managing turfgrass stands that are free of damaging turf pests is essential to providing aesthetically pleasing lawns, golf courses, and sports fields. Creeping bentgrass is one of the most common turfgrass types found on golf course putting greens but is also used on golf course fairways and tee boxes. There are many diseases that can be found on creeping bentgrass when environmental conditions are favorable. Of these diseases, dollar spot is the most common. When dollar spot is present, half-dollar sized spots of bleached turf can be seen. In order to prevent these easily noticeable spots from appearing, fungicide applications are required in a given growing season to prevent the pathogen from infecting. Available fungicides are very effective at providing control but can be very costly. Beyond fungicide use, other research has shown various cultural practices to reduce disease incidence. Previous research has shown that iron sulfate applied to creeping bentgrass can reduce dollar spot epidemics without the use of fungicides. Laboratory studies have shown a similar trend as ferrous sulfate at varying concentrations directly suppressed dollar spot pathogen growth. In both cases, a limited range of ferrous sulfate rates was tested. To obtain more information we explored use rates of ferrous sulfate required to suppress 50% of dollar spot in the field and in vitro. Ferrous sulfate rates in field trials ranged from 0 to 97.6 kg ha⁻¹ . Results from these trials were used to create a hyperbolic regression. Using this model, we were then able to determine that 26.4 kg ha⁻¹ iron sulfate was required to suppress 50% of the dollar spot in the field. For the laboratory studies we concluded that ferrous sulfate suppresses 50% of the dollar spot pathogen mycelial growth between 480 and 720 mg L ⁻¹ iron sulfate concentrated potato dextrose agar. Although there are many different fungicides available for dollar spot control, the active ingredient chlorothalonil is the most common used. Due to the mode of action which chlorothalonil exhibits, it is much less likely that the pathogen causing dollar spot can become resistant. Although resistance is not an issue, governmental annual site-use limitations restrict turf managers from achieving desirable control. The use of iron sulfate in conjunction with chlorothalonil has not been previously studied. Chlorothalonil treatments were applied at a range of labeled use rates across plots treated with and without 48.8 kg ha⁻¹ ferrous sulfate applied bi-weekly. Ferrous sulfate reduced the chlorothalonil rates necessary for disease reduction. If a threshold of 80% is used, up to 50% reduction in chlorothalonil use was observed. Supplemental studies investigating the duration of control achieved by the combination showed an increase of up to 5 days and eliminated the need for two applications across one season. This research fills a huge gap in our knowledge base on the practical use of iron sulfate for dollar spot control.

Parasitic weeds of the family Orobanchaceae attach to the roots of host plants via haustoria capable of drawing nutrients from host vascular tissue. Species in this family span the spectrum of host nutrient dependency, allowing comparisons that provide insight into parasite adaptation. A key aspect of this is the relationship between parasite metabolism and the metabolite profile of its host. To what extent does the metabolite profile of the parasite depend on that of the host? Do parasites that differ in host-dependency also differ in their metabolism or do they use common metabolic strategies? These questions were addressed using comparative profiling of primary metabolites to gain insight into carbon and nitrogen assimilation by the obligate holoparasite Phelipanche aegyptiaca and the facultative hemiparasite Triphysaria versicolor. First, metabolite profiles of these parasites and their hosts were compared during the key life stages before and after haustorial attachment. Second, the impact of specific variations in host metabolism was analyzed for P. aegyptiaca growing on Arabidopsis thaliana hosts that had mutations in amino acid metabolism but otherwise identical genetic backgrounds. Comparison of P. aegyptiaca and T. versicolor metabolite profiles identified substantial differences in the stages spanning the transition from pre-haustorial development through post-haustorial feeding. Each parasite species is distinct from the other and from their hosts. For parasites growing on host lines that differ in amino acid content, the size of P. aegyptiaca tubercles decreased when grown on the aap6 mutant line, which has decreased levels of asparagine in the phloem sap compared to the wild type. However, altered amino acid levels in other lines did not impact P. aegyptiaca growth, indicating that this parasite has ability to compensate for variation in host metabolic composition. This research highlights the importance of aspartate and asparagine to early post-attachment metabolism in both P. aegyptiaca and T. versicolor and through host deficiencies possibly associated with decreased growth in P. aegyptiaca. Overall, this work provides insights both into the metabolism of parasitic plants and lays the foundation for the development of new metabolism-based control strategies. Ph. D. Parasitic weeds of the plant family Orobanchaceae attach to the roots of host plants via haustoria. Parasite haustoria embed into the host plant and are capable of drawing nutrients from host vascular tissue. Species in this family span the spectrum of the extent to which a parasitic plant may depend on its host for nutrients. This allows comparisons that provide insight into the ways in which parasites adapt. A key aspect of this is the relationship between the metabolite profile of the parasite and the metabolite profile of the host. To what extent does the metabolite profile of the parasite depend on that of the host? Do parasites that differ in host-dependency also differ in their metabolism or do they use common metabolic strategies? These questions were addressed using comparative profiling of primary metabolites to gain insight into carbon and nitrogen assimilation by the obligate parasite Phelipanche aegyptiaca (which cannot perform photosynthesis) and the facultative parasite Triphysaria versicolor (which can perform photosynthesis). First, metabolite profiles of these parasites and their hosts were compared during the key life stages before and after haustorial attachment. Second, the impact of specific variations in host metabolism was analyzed for P. aegyptiaca growing on Arabidopsis thaliana hosts. These hosts had mutations in enzymes related to amino acid metabolism but otherwise identical genetic backgrounds. Comparison of P. aegyptiaca and T. versicolor metabolite profiles identified substantial differences in the stages spanning the transition from pre-haustorial development through post-haustorial feeding. Each parasite species is distinct from the other and from their hosts. For parasites growing on host lines that differ in amino acid content, the size of P. aegyptiaca tubercles decreased when grown on the aap6 mutant line, which has decreased levels of asparagine in the phloem sap compared to the wild type. However, altered amino acid levels in other lines did not impact P. aegyptiaca growth, indicating that this parasite has ability to compensate for variation in host metabolic composition. Overall, this work provides insights both into the metabolism of parasitic plants and lays the foundation for the development of new metabolism-based control strategies.

The interaction between Arabidopsis and its natural downy mildew pathogen, Hyaloperonospora arabidopsidis (Hpa), provides a model for understanding how oomycetes colonize plants. Hpa is a model organism for many highly destructive oomycete pathogens and transcriptomics of this interaction have been well-documented. However, the material in these studies has been derived from infected leaves that contain a mix of pathogen-proximal and pathogen-distal plant cells. The most direct interactions between Arabidopsis and Hyaloperonospora arabidopsidis occur in haustoriated cells- where the pathogen can secrete effectors and acquire nutrients needed for successful colonization and reproduction. These cells are difficult to isolate due to their limited number and ephemeral nature. I have developed a method to isolate the translatome (i.e., mRNAs associated with ribosomes) of pathogen-proximal cells. This method utilizes translating ribosome immuno-purification technology (TRAP), regulated by both pathogen-responsive and tissue-specific promoters, to isolate mRNAs that are being translated in pathogen-proximal cells. Compared to "bulk" transcriptomics of material isolated from homogenized leaves, this method will enrich for transcripts that are differentially expressed, and translated, in pathogen-proximal cells. From this method, RNA was isolated in amount and quality sufficient for sequencing. This sequencing data will enable the discovery of plant genes that may be manipulated by the pathogen to suppress defense responses and extract nutrients. Ph. D. The interactions between plants and the pathogens that feed on them are complex and at times difficult to study. Among the many different types of plant pathogens, oomycetes (a class of fungus-like organisms) are especially destructive. Using Arabidopsis and its natural downy mildew pathogen, Hyaloperonospora arabidopsidis (Hpa) as model for understanding how oomycetes colonize plants, I hope to learn more about plant-pathogen interactions. Hpa is a model organism for many highly destructive oomycete pathogens and several aspects of this interaction have been well-documented. However, the material in these studies has been derived from infected leaves that contain a mix of plant cells that are both in direct contact with the pathogen, or from uninfected areas of the plant. The most direct interactions between Arabidopsis and Hpa occur in cells that have been invaginated with a pathogen feeding structure called a haustorium. These cells are difficult to isolate due to their limited number and ephemeral nature. I have developed a method to isolate the translatome (i.e., mRNAs that are being translated by and are associated with ribosomes) of pathogen-proximal cells. This method utilizes translating ribosome immuno-purification technology (TRAP), regulated by both pathogen-responsive and tissue-specific promoters, to isolate mRNAs that are being translated in pathogen-proximal cells. Compared to “bulk” transcriptomics of material isolated from homogenized leaves, this method will enrich for transcripts that are differentially expressed, and translated, in pathogen-proximal cells. From this method, RNA was isolated in amount and quality sufficient for sequencing. This sequencing data will enable the discovery of plant genes that may be manipulated by the pathogen to suppress defense responses and extract nutrients.

Crown gall of grapevine (Vitis spp.) is a serious and economically important disease caused by the bacterial pathogen Rhizobium vitis, which transforms healthy plant cell genomes leading to hypertrophic and hyperplastic growth of affected plant cells. Recent studies have documented a strong inhibitory effect against Japanese tumorigenic R. vitis isolates by a newly identified non-tumorigenic strain of R. vitis, ARK-1. We conducted co-inoculation assays in tomato (Solanum lycopersicum) and wine grape cultivars (Vitis vinifera) with four tumorigenic isolates of R. vitis from Virginia. These tumorigenic isolates were co-inoculated with ARK-1 in various ratios and resulting gall incidence and gall size were measured. Analysis was conducted with the generalized linear mixed model (GLIMMIX) in SAS (ver. 9.4). ARK-1 significantly reduced both the mean probability of gall formation and the mean gall size (P < 0.05). ARK-1 efficacy against combinations of two or four tumorigenic isolates and up to twice as many cells of tumorigenic isolates was also significant. However, there was an indication of a loss of efficacy when ARK-1 was challenged with four isolates at four times the cell number of ARK-1. Also, the efficacy of ARK-1 was influenced by both the specific isolate and host plant used in the study. Our results suggest that ARK-1 has promising potential as an effective biological control agent for grapevine crown gall in the United States. MSLFS Crown gall of grapevine (Vitis species) is a serious and economically important disease to the grape production industry caused by the bacterium Rhizobium vitis. This pathogen alters the genetic material of plant cells leading to cancer-like growth of affected plant cells (tumors); which may lead to the death of a grapevine. Recently, a non-tumor inducing strain of R. vitis (ARK-1) was documented to inhibit gall induction by tumor-inducing R. vitis strains in Japan. To see if ARK-1 would be a good candidate for biological control of the tumor-causing strains of R. vitis in the US, we tested it against four tumor-inducing strains in both tomato (Solanum lycopersicum) and grapevine (Vitis vinifera). The tumor-inducing strains were mixed with ARK-1, injected into plant stems in various ratios, and resulting gall incidence and gall size were measured 42 to 90 days later. ARK-1 significantly reduced both the probability of gall formation and the gall size in both tomatoes and grapevines. ARK-1 was an effective agent against mixtures of one, two, or four tumor-inducing strains and provided control even when outnumbered two to one by cells of tumor-inducing strains. The efficacy of ARK-1 was influenced by the specific strain, relative cell number to tumor-inducing strains, and host plant used in the study. Our results suggest that ARK-1 has promising potential as an effective biological control agent for grapevine crown gall in the United States.

Regalia®, an extract from giant knotweed (Reynoutria sachalinensis (REYSA)), is an organic immune system response (ISR) treatment that can mitigate fungal and bacterial infections by stimulating the genetic immune response system of a plant. The mode-of-action of Regalia® is to penetrate the plant tissue and stimulate a biodynamic response to resist pathogens. The primary response to Regalia® is the production of reactive oxygen species (ROS) that disable pathogens, and a thickening of cell walls to inhibit pathogen penetration. Regalia® is an uncommon fungicide since the protection is provided by the plant itself and not directly by the agent, and because the activity is internal rather than external. This process provides protection from pathogens that get past surface protectants. Because ISR agents require plant resources to be effective, there is a risk that they can disrupt the metabolic processes of the plant, or there can be an allocation cost that could diminish the plant vigor. This study examines if the ISR activity of Regalia® presents any phytoxicity or impact to vigor in four varieties of cider apples (Malus ×domestica cvs. Dabinett, Medaille d’Or, Chisel Jersey, and Redfield). This quantitative study, based on measured metrics of plant growth during the study period, showed that Regalia, applied with Cueva or Serenade, had no effect on primary or secondary shoot length or the number of shoot leaves on any of the four test varieties. MALS

Spring dead spot (SDS) is the most destructive disease of bermudagrass (Cynodon spp.) in Virginia. SDS infects bermudagrass in the fall with symptoms appearing in the spring when dormancy breaks. Patches are sporadically distributed but generally reoccur in the same location. Chemical control options are expensive with inconsistent results. Our objectives were to develop SDS incidence maps, investigate methods to analyze these maps, and evaluate suppression efficacy of incidence-map-based chemical applications. Methods were developed to build SDS incidence maps in 2016 and 2017. 2016 SDS incidence maps were compared for spatial accuracy to Digital Orthophoto Quarter Quadrangle (DOQQ), ground-validated differential GPS coordinates, and to 2017 SDS incidence maps, with average deviations of 1.3 m, 1.6 m, and 0.1 m, respectively. Digital Image Analysis (DIA) of aerial maps was compared to a point-intersect method for validation with a significant linear relationship (r2 = 0.77, P ≤ 0.0001). In the fall of 2016 and 2017, a site-specific penthiopyrad (SSP) treatment was evaluated against blanket, full-coverage applications of penthiopyrad (BP) and tebuconazole (BT), and an untreated control. Treatments were compared using DIA, post-treatment SDS patch count (PC), and SDS patch reduction (PR). Across all three metrics, the penthiopyrad treatments were statistically superior to both the tebuconazole and untreated. SSP compared favorably to BP for DIA, but BP had 2.57 fewer PC (LSD = 2.05) and a greater PR by 2.58 (LSD = 2.55). SSP using SDS incidence maps required 51% less fungicides in 2016 and 65% less in 2017 when compared to BP. Master of Science in Life Sciences