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Browsing by Author "Roberts, Robyn, committee member"

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    An NLR gene likely underlying RMES1 provides global sorghum resistance bolstered by RMES2
    (Colorado State University. Libraries, 2023) VanGessel, Carl, author; Morris, Geoffrey, advisor; Nalam, Vamsi, committee member; Roberts, Robyn, committee member; Mason, Esten, committee member
    Breeding for aphid host plant resistance in sorghum has been an area of interest since the emergence of Melanaphis sorghi in North America a decade ago. In order to develop durable sorghum aphid resistance, breeders must be equipped with tools (trait package) and knowledge (molecular mechanisms) of host plant resistance. In this dissertation, I characterize the current state of sorghum aphid breeding and propose a genotype to phenotype map for the major source of global resistance, Resistance to Melanaphis sorghi 1. Relying on near-isogenic lines, I demonstrate that RMES1 is applying selection pressure to sorghum aphid through reduction in fecundity that discriminates among aphid species. In global sorghum lines, RMES1 is rare whereas a second resistance source, RMES2, is common and present in historic breeding germplasm. I mapped RMES2 in Haitian breeding populations where it contributes fitness increases while lacking antagonistic pleiotropy and is selected for alongside RMES1. These results suggest breeding programs may unknowingly be deploying both sources of resistance which in combination are reducing the likelihood of M. sorghi biotype shifts to overcome RMES1. As aphid resistance may rely on phytochemical and/or induction with extended phenotypes regarding aphid populations, I used pan-genomic, transcriptomic, and metabolomic resources to describe the molecular mechanism of RMES1. Structural variation at the Chr06 locus underlies presence/absence variation of several nucleotide-binding leucine-rich repeat receptor (NLR) genes. Two of these candidate genes, SbPI276837.06G016400 and SbPI276837.06G016600, are representatives of two orthologous NLR groups which have genomic and transcriptomic evidence of underlying RMES1 resistance. The PAL branch of the salicylic acid pathway is the primary phytohormone pathway responsible for RMES1-induced resistance. Finally, metabolome reorganization mirroring transcriptome changes suggest RMES1 is inducing multiple downstream mechanisms responsible for reducing aphid fecundity. While the causal gene underlying RMES1 remains to be cloned and the eliciting aphid factor is unknown, this research suggests that gene-for-gene dynamics could lead to resistance-breaking biotype shifts and combining RMES1 with additional resistance genes e.g. RMES2, will help achieve durability.
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    Exploring the hemp virome and assessing hemp germplasm for resistance to emerging pathogens
    (Colorado State University. Libraries, 2024) Hackenberg, Laine, author; Nachappa, Punya, advisor; Roberts, Robyn, committee member; Stenglein, Mark, committee member
    Hemp (Cannabis sativa L.), commonly grown for its seeds, fiber, and non-psychoactive cannabinoids, has been experiencing a resurgence in the United States with its recent legalization. While farmers across the nation have readily adopted this crop, resources for pest management are still lacking, particularly regarding the diversity and distribution of pathogens. As production increases and the crop diversifies, the emergence and spread of these pathogens are certain. To circumvent loss due to disease, research is needed understand the threats and to identify sustainable management options. The goal of this study is to describe the diversity and distribution of viruses/viroids infecting hemp in Colorado and to determine if there is genetic resistance to pathogens in hemp. The objectives of this study are to 1) characterize the virome of different hemp cultivars throughout the growing season across different locations and 2) screen a panel of genetically unique genotypes of hemp for resistance to emerging viruses/viroids of hemp. Throughout 2021 and 2022, the hemp virome was examined in four major hemp producing regions of Colorado. In total, nine fields were sampled, and each field was visited during three phenological stages (early vegetative, late vegetative, and mature flowering) in order to characterize the virome throughout the growing season. Leaf tissue samples were collected from two cultivars of hemp from each field site. These tissue samples were submitted for High Throughput Sequencing (HTS) and upon bioinformatic analysis, candidate virus/viroid sequences were validated. Across both years, a total of seven viruses were identified: Alfalfa mosaic virus (AMV), Beet curly top virus (BCTV), Cannabis cryptic virus (CanCV), Cannabis sativa mitovirus (CasaMV1), Grapevine line pattern virus (GLPV), Opuntia umbra-like virus (OULV), and Tomato bushy stunt virus (TBSV). All viruses identified had >97% nucleotide identity to the nearest GenBank accession. Between individual cultivars isolated from the same field, both similar and unique viromes were observed. Viral diversity and incidence increased as the growing season progressed for both years. The three viruses that were most commonly found across all regions were CasaMV1, GLPV, and BCTV. Dominating the virome in viral load were CasaMV1 and GLPV. Given the prevalence of BCTV in the virome, in addition to its prevalence in hemp across the western United States, 13 genotypes of hemp were screened for resistance to this pathogen. These genotypes of hemp are genetically diverse, which will aide in the discovery of candidate genes involved with resistance. BCTV is the causal agent of curly top disease which can have drastic symptomology in hemp plants, causing malformed growth, stunted plants, and crop loss up to 100%. Varying BCTV copy number was observed across the hemp genotypes. Additionally, percent disease index (PDI) was analyzed to determine the frequency of infection of individual genotypes. Two of the genotypes were observed to have a lower PDI than the others, 4587 and 4710. Hop latent viroid (HLVd) has been emerging as a threat to the cannabis industry. It has been described across North America but is believed to be worldwide due to its global distribution in hops. HLVd has been documented to cause drastic reduction in cannabinoid content in mature inflorescences and therefore has the potential for substantial economic losses. Although not identified within the 2021 or 2022 virome, HLVd was determined to be an important threat facing hemp production therefore it was included in the screening. Similarly to BCTV, a panel of 14 genetically unique genotypes of hemp were analyzed for resistance to HLVd. Resistance was identified in a single genotype, 517, which had a lower frequency of infection than the others. However, no varying viroidal loads were observed between genotypes. Throughout this study, viruses associated with hemp were described as well as the identification of genetic resistance to emerging pathogens. This work will help to further integrated pest management strategies and promote sustainable agriculture.
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    HopBA1, a pathogen virulence factor, reveals tissue-specific immune responses within the Pseudomonas syringae pv. tomato–Nicotiana benthamiana pathosystem
    (Colorado State University. Libraries, 2024) Todd, Tyler Scott, author; Nishimura, Marc, advisor; Sloan, Dan, committee member; Roberts, Robyn, committee member
    Plant pathogens represent a major threat to food security as they dramatically reduce crop yield, impact the expression of desirable traits, and reduce post-harvest longevity. To infect host plants, bacteria like Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) utilize a type III secretion system to deliver virulence proteins (also known as effectors) into the host cytoplasm to suppress immunity. In response, plants have evolved intracellular immune receptors that perceive immunosuppression and reactivate immunity. Thus, a given pathogen effector can both suppress and activate immunity depending on the host genome. Dissecting the molecular mechanisms of plant pathogen effectors helps inform our understanding of both disease and resistance. The present work reveals an uncharacterized role for Pst DC3000 as an aggressive vascular pathogen, causing systemic infection in the model plant Nicotiana benthamiana (Nb). Further, it establishes that the bacterial effector HopBA1 inhibits movement through the vascular system, despite increasing pathogen persistence within the primary infection site in leaves. Simultaneously, HopBA1 was found to induce irreversible upward vertical bending (i.e., hyponasty) in the petioles of infiltrated leaves, a novel phenotype for a bacterial effector. LC-MS/MS and RNA-Seq revealed phytohormone alteration (notably, a reduction in auxin and jasmonic acid-related metabolites) and transcriptional reprogramming of both developmental and defense genes. HopBA1-dependent growth restriction was suppressed in Nb eds1 (ENHANCED DISEASE SUSCEPTIBILITY 1) mutant plants, which still undergo HopBA1-induced hyponasty. Together, these results suggest that (1) HopBA1 triggers tissue-specific immune responses and (2) hyponasty is due to HopBA1's virulence activity, rather than host immune activation. Thus, HopBA1 in combination with the model pathogen Pst DC3000 becomes an important tool to dissect the poorly understood area of vascular-specific immunity. Vascular pathogens are particularly devastating and difficult to manage in crop species owing to the pathogen's internal location and systemic route of infection, making this research useful in crop improvement.
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    TALE-bound QTL: a computational investigation of bacterial effector association with resistance quantitative trait loci in Oryza sativa
    (Colorado State University. Libraries, 2022) Sharkey, Jacob Emmett, author; Leach, Jan E., advisor; Huerta, Alejandra I., committee member; Nishimura, Marc, committee member; Roberts, Robyn, committee member
    Durable resistance to Xanthomonas oryzae pathovars oryzae (Xoo) and oryzicola (Xoc), which cause bacterial blight and bacterial leaf streak, respectively, is highly sought after in rice (Oryza sativa) due to the pathogens ability to impact maximum attainable yields. Regions of the rice genome associated with quantitative resistance to multiple strains of Xoo and Xoc, known as quantitative trait loci (QTL), were previously identified using a multi-parent advanced generation intercross (MAGIC) rice population and a combination of genome wide association studies and interval mapping. These QTL have been associated with decreased lesion lengths by Xoc and Xoo on rice. What remains unknown is the molecular basis for the induction of genes under these QTL during pathogen infection. Considering our biological question "what is the molecular basis for regulation of resistance QTL associated with Xoo and Xoc?", we predicted that part of the answer could be found by investigating the bacteria's direct interaction with the O. sativa genome. Upon infection, Xoo and Xoc injects the host with DNA-binding TALE (transcription activator-like effector) proteins. These effectors, when bound to their target plant gene promoter, induce gene transcription. We hypothesize that differential interactions of TALE with promoters of rice genes under the QTL lead to the resistant/susceptible phenotypes exhibited across varieties. To test this, we designed a pipeline that predicts TALE-regulated candidate genes involved in quantitative resistance. This pipeline identifies genes that meet three criteria: (1) the presence of a binding site for an X. oryzae TALE in the gene's promoter, a strong correlation between binding site presence, and disease phenotypes and overlap of the gene with a resistance QTL. We used this pipeline with genomic and phenotypic data for the eight MAGIC founders to identify candidate genes involved in resistance against seven Xoo and Xoc strains. Candidate genes identified include ones encoding a patatin-like phospholipase and multiple NB-ARC containing proteins such as the Mla1 protein. Here, we exploit the abundant genomic data for the rice-X. oryzae systems and the ability to predict direct associations between bacterial proteins and plant genomes, to propose a method that could streamline the identification of genes involved in quantitative resistance to TALE- harboring Xanthomonas.