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Browsing by Author "Nachappa, Punya, committee member"

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    Characterizing host genetic resistance to Wheat streak mosaic virus (WSMV) and Fusarium wilt disease
    (Colorado State University. Libraries, 2021) Xie, Yucong, author; Pearce, Stephen, advisor; Argueso, Cristiana, committee member; Muñoz-Amatriaín, María, committee member; Nachappa, Punya, committee member
    Crop production is limited by a variety of biotic stresses caused by pathogens. This study focuses on wheat streak mosaic disease in wheat, caused by the viral pathogen Wheat streak mosaic virus (WSMV), and Fusarium wilt disease in banana, caused by the fungal pathogen Fusarium oxysporium f.sp. cubense (Foc). In this dissertation, I applied genomic and transcriptomic tools to study the Wsm2 locus that confers genetic resistance to WSMV. Analyzing exome and transcriptome reads from wheat lines carrying Wsm2, I characterized structural variations and identified unique transcripts specific to these Wsm2 carrying lines. Moreover, examination of candidate genes within the Wsm2 interval identified several tandemly duplicated candidate genes annotated as Bowman-Birk inhibitor (BBIs), which triggered my interests to perform a genome-wide characterization of this gene family in wheat. I studied the possible mechanisms behind its copy number and functional domain duplications and analyzed its diverse role in plant biotic and abiotic stress using wheat RNA-seq expression data. Finally, I analyzed a time course transcriptomic dataset from banana root infected with Foc subtropical race 4 strain (Foc-STR4). I used gene co-expression assembly network (WGCNA) to study host plant transcriptional response to Foc infection and analyzed the expression profiles of candidate genes underlying a novel locus conferring resistance to Foc-STR4 and prioritized candidates. In summary, this dissertation studied genetic variants underlying host genetic resistance to WSMV and Foc and shed light on plant defense mechanisms against these two important crop pathogens.
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    Developing integrated pest management tactics for alfalfa mosaic virus and its aphid vector in chile peppers
    (Colorado State University. Libraries, 2023) Janecek, Taylor, author; Szczepaniec, Adrianna, advisor; Nachappa, Punya, committee member; Uchanski, Mark, committee member
    Alfalfa mosaic virus (AMV, Bromoviridae: Alfamovirus) is a virus transmitted to plants by aphids in a non-persistent manner. The virus was first identified in chile peppers Capsicum annuum L. (Solanales: Solanaceae) in Southern Colorado in 2019. The goal of this research was to explore management strategies to suppress the virus given its devastating impact on the yield and quality of chile peppers. The objectives were to: 1) determine whether chile peppers have innate resistance to AMV, 2) test the effectiveness of host plant resistance and planting date modifications to suppress the virus in the field, 3) determine whether AMV is seed transmissible, and 4) survey abundance and diversity of aphids (Hemiptera: Aphididae) that likely transmit AMV in the system. In the greenhouse experiment, I found significant differences among varieties of chile peppers in the severity of AMV symptoms and identified a variety suitable for a field experiment. In the field, I found that the susceptible variety, Joe Parker, which tended to have high AMV symptoms in the greenhouse, was also highly susceptible to AMV in the field. Conversely, Mira Sol, which appeared to have resistance to the virus in the greenhouse screening assay had low incidence of AMV symptoms and low AMV titers in the field as well. Planting date also played an important role in symptom severity, where late planted peppers (mid-June) had significantly lower severity of AMV symptoms than peppers planted at conventional and early planting dates (the middle and end of May). Despite this, the yield and quality of peppers planted early was significantly greater than that of peppers planted later in the season. In addition, there was evidence of seed transmission of AMV in chile peppers, with 10% for Mira Sol and 2% for Joe Parker, from seeds collected from infected peppers had AMV. Lastly, I found high diversity of aphid species within my experimental plots (14-16 species) and lower diversity in nearby alfalfa fields (4-5 species). Moreover, severity and titers of AMV were positively correlated with earlier planting date, which was likely related to higher aphid densities early in the season. This research contributed to formulating integrated tactics that chile pepper producers can implement in their production to suppress the impact of AMV on the crop. Finally, this is the first report of AMV transmission through seed in peppers and is the first study describing this pathosystems in Colorado.
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    High throughput characterization of bunyavirus diversity, ecology, and reassortment potential
    (Colorado State University. Libraries, 2023) Kapuscinski, Marylee, author; Stenglein, Mark, advisor; Kading, Rebekah, committee member; Wilusz, Jeff, committee member; Ebel, Greg, committee member; Nachappa, Punya, committee member
    Bunyavirales is an important group of viral pathogens with significant economic impacts. The Bunyavirales order contains the largest number of RNA viruses and can cause disease in plants, animals, and humans [1]. Notable plant pathogens include Tomato Spotted Wilt virus [2] which results in significant agricultural losses. Notable animal pathogens include Rift Valley fever virus [3] and Schmallenberg [4] virus resulting in significant livestock loss. Notable human pathogens include Crimean-Congo hemorrhagic fever virus [5], hantavirus [6], and La Crosse virus [7]. The effects of bunyavirus infections are felt worldwide because bunyaviruses are distributed globally. The emergence of novel bunyaviruses continues to threaten agricultural and livestock industries as well as human health. Where or when a novel bunyavirus might emerge is unknown. Predicting emergence is difficult for three reasons. First, because bunyaviruses are RNA viruses [8], the high error-rate of the RNA-dependent RNA polymerase results in a large genetic diversity within a population of viruses [9]. Secondly, many important bunyaviruses are arthropod-borne [10], resulting in an intricate lifecycle between an invertebrate and vertebrate host. This results in constantly changing genetic diversity due to different selective pressures from different host types [11]. It also results in an ever-expanding geographic range as vector range expands due to climate change and vertebrate host range changes due to urbanization, industrialization, and deforestation [12–16]. Third, bunyaviruses have segmented genomes which allows them to reassort and produce viral progeny with an altered vector-host range, pathogenesis, and virulence [1,17,18]. Therefore, this body of work aims to increase our ability to understand bunyavirus emergence and reassortment potential as a way to aid in outbreak preparedness and early response systems. To do this, we've combined traditional surveillance data with modern bioinformatics to expand our knowledge of bunyavirus genetic diversity, ecology, and reassortment potential. Using whole-genome sequencing, we've characterized the genomes of 99 bunyaviruses, some of which have never been sequenced before. This aids in our understanding of the genetic diversity, co-infection dynamics, and reassortment potential. Next, we used existing metadata from orthobunyavirus sequences to determine reassortment potential given a shared geographic and vector-host range. Finally, we've developed a novel molecular assay to evaluate reassortment potential based on replication and transcription compatibility. Together, we've combined the strengths of viral surveillance and modern bioinformatics to demonstrate the benefit of combining both. We've developed systems that will help to delineate the mechanisms that either promote or inhibit reassortment potential, ultimately aiding in early response systems for outbreak preparedness.
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    Life cycle environmental impacts of utilizing hemp seed meal as a protein source in sheep feedlot rations
    (Colorado State University. Libraries, 2023) Clark, Samantha Maye, author; Dillon, Jasmine, advisor; Archibeque, Shawn, committee member; Nachappa, Punya, committee member; DiVerdi, Joseph, committee member
    Hemp seed meal is a protein-rich byproduct of the hemp industry, obtained from the cold press extraction process used to produce hemp oil. The objective of this work was to evaluate the environmental impact of using hemp seed meal as a protein supplement in sheep production. A cradle-to-gate life cycle assessment (LCA) was conducted on three sheep production systems which differed in the feedlot phase: one fed a feedlot ration containing soybean meal as the protein source (soybean meal diet), one fed hemp seed meal in the feedlot ration (hemp diet), and one fed organic hemp seed meal in the feedlot ration (organic hemp diet). Animal performance data were collected from a nutrition trial. Hemp production, harvest, and processing data were provided by a hemp product company. Economic and physical allocation were applied to the hemp diet systems, and the ReCiPe Midpoint (H) methodology was used to calculate the global warming (i.e., carbon footprint), water consumption, land use, and fossil resource scarcity impacts on a per kg lamb live weight basis for each system. Carbon footprint ranged from 10.1 to 11.4 kg CO2eq/kg LW, water consumption ranged from 1.3 to 4.2 m3/kg LW, fossil resource scarcity ranged from 0.5 to 0.8 kg oil eq/kg LW, and land use ranged from 2.8 to 6 m2a crop eq/kg LW. Impact assessment results were not sensitive to a 10 or 20% increase in electricity demand at processing. The use of IPCC Tier 2 methods for estimating enteric methane emissions from sheep resulted in a 7.5–8.5% increase in the carbon footprint, relative to a mechanistic equation present in the Ruminant Nutrition System model. Physical allocation resulted in greater impacts of the hemp diet systems than the soybean diet systems for all categories except land use. However, economic allocation resulted in greater impacts for the soybean diet systems than the hemp diet systems for all categories evaluated. This was explained by inherent differences between the allocation method, as physical allocation attributed 80% of the environmental burden to hemp seed meal, while economic allocation attributed 0% of the environmental burden to hemp seed meal due to the current lack of an economic value for hemp seed meal. The production volume of dependent products ("dependent products", or products for which a change in demand does not affect production volume, commonly referred to as co- or byproducts) are driven by monetary value of the determining product (the product for which a change in demand affects the production volume), but relationships between co-products change overtime. Therefore, as the hemp industry continues to develop, an economic value may be placed on hemp seed meal with implications for its relative ability to reduce the environmental impacts of livestock production. As agricultural industries strive to become more environmentally efficient, they must be adaptive to changes in both monetary value and environmental impact, which are intrinsically related. This research demonstrated the importance of allocation choice in assessing the impact of feeding byproducts on the environmental impact of livestock production systems. Economic allocation better reflected the monetary driving factor for hemp production than physical allocation. As such, the inclusion of hemp seed meal in a feedlot ration reduced the environmental impact of sheep production systems.
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    Rangeland management impacts on native bee diversity and pollination services in the shortgrass steppe of Colorado's Front Range
    (Colorado State University. Libraries, 2021) Thapa-Magar, Khum Bahadur, author; Davis, Thomas Seth, advisor; Fernández-Giménez, María E., committee member; Laughlin, Daniel, committee member; Nachappa, Punya, committee member
    To view the abstract, please see the full text of the document.
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    Reframing viral infections as acute metabolic disorders: dengue viruses and their dependency on host metabolic pathways
    (Colorado State University. Libraries, 2022) St. Clair, Laura A., author; Perera, Rushika, advisor; Belisle, John, committee member; Nachappa, Punya, committee member; Wilusz, Jeff, committee member; Zabel, Mark, committee member
    Dengue viruses (DENVs) are the etiological agent of the world's most aggressive arthropod-borne disease. At present, there are no available antivirals against DENVs. This fact underscores a dire need to examine host-virus interactions to identify and develop novel therapeutic approaches. As obligate intracellular parasites, DENVs are reliant upon and hijack several host metabolic pathways both to fulfill their replicative needs, and to evade the host immune response. We and others have previously established that infection with DENVs causes significant perturbation to host lipid metabolism, including elevations in sphingolipids in both the human and mosquito host. In addition, we and others previously discovered that the DENV NS1 protein increases sialidase activity in both in vitro and in vivo models leading to increased endothelial hyperpermeability and vascular leakage which are hallmarks of severe dengue. To further clarify and characterize these previous works, we have performed siRNA-mediated loss of function studies using human hepatoma cells (Huh7 cells) on several metabolic pathways altered during DENV2 infection. First, we examined the role of acyl-CoA thioesterases, enzymes responsible for controlling the intracellular balance of activated fatty acids and free fatty acids, on the DENV2 lifecycle. In these analyses, we determined that the cytosolic ACOT1 enzyme had an inhibitory effect on DENV2 replication and release, while mitochondrial ACOT (ACOTs 2 and 7) functionality was critical for viral replication and release. Moreover, we identified several enzymes within the ACOT family whose expression was dependent on ACOT2 and ACOT7 expression. These results highlighted complex relationships between ACOTs and DENVs, as well as identified yet unknown functional interdependence between ACOT enzymes. Next, we expanded our previous understanding of the relationship between DENVs and the human sialidase enzymes (NEU1-4). While previously studies linked upregulation of these enzymes with DENV2 pathology, we provide the first evidence showing that NEU1-4 functionality is vital for DENV2 genome replication and viral egress. Moreover, our analyses also revealed previously unknown functionality of NEU4 or its downstream products as transcriptional regulators for NEU1-3. Finally, we provide the first profile of the effect of loss of function of enzymes within the entire sphingolipid metabolic pathway (as identified through KEGG pathway database) on the DENV2 life cycle. In this study, we identified that enzymes involved the sphingomyelinase and salvage pathways of ceramide synthesis as opposed to de novo ceramide synthesis were critical to DENV2 release from Huh7 cells. In addition, we determined that enzymes involved in the synthesis and degradation of glycosphingolipids were vital for DENV2 release. An especially intriguing result within this arm of sphingolipid metabolism was that the two enzymes which hydrolyze GluCer had differential effects on DENV2 replication and release. GBA1 (lysosomal) had an antiviral effect on DENV2, while GBA2 (non-lysosomal) was required for DENV2 replication and release. This prompted us to profile the changes that occur to glycosphingolipids (GSLs) during infection, and we uncovered several species of GSLs that are elevated during infection. Moreover, we identified that Ambroxol HCl, a pharmaceutical GBA1 chaperone/GBA2 inhibitor, was able to abrogate these elevations in GSLs. Combined, our results allowed us to propose a novel function for GBA2 as a GluCer recycling enzyme during DENV2 infection. In conclusion, together, the work in this dissertation highlights critical metabolic nodes that impact virus replication and provides new directions for investigating viral infections as acute metabolic diseases.