Molecular studies on the sweet potato virus disease and its two causal agents

The studies presented in this thesis contribute to an increased understanding of the molecular aspects, variability and interaction of the two most important viral pathogens of sweet potato (Ipomoea batatas L): Sweet potato feathery mottle virus (SPFMV) and Sweet potato chlorotic stunt virus (SPCSV), which cause the severe sweet potato virus disease (SPVD) when co-infecting sweet potato plants. SPVD is the most important disease affecting sweet potato in Africa, and may be the most important virus disease of sweet potato globally. The coat protein gene sequences of several African SPFMV isolates were determined and compared by phylogenetic analyses. Results showed that East African SPFMV isolates were genetically distinct. They could furthermore be divided into two serotypes which differed in their ability to systemically infect the sweet potato cultivar Tanzania. The aetiology of SPVD was studied in sweet potato plants co-infected with SPFMV and SPCSV using nucleic acid hybridisation, bioassays, tissue printing and thin section immunohistochemistry. Resistance to SPFMV in East African sweet potato cultivars was found to be due to inhibition of virus replication rather than movement and resistance was suppressed by infection with SPCSV, resulting in a ca. 600-fold increase in titres of SPFMV. Furthermore, in SPVD affected plants SPFMV is detected outside of the phloem, whereas SPCSV is detected only inside the phloem, which suggests novel as yet unknown mechanisms how SPCSV synergises SPFMV. The genomic sequence of SPCSV was determined. It was composed of two RNA molecules (9407 and 8223 nucleotides), representing the second largest (+)ssRNA genome of plant viruses. The genomic organization of SPCSV revealed novel features for the genus Crinivirus, such as i) the presence of a gene putatively encoding an ribonuclease III-like protein, ii) near-identical, 208 nucleotides long 3’-sequences on both viral RNAs, and iii) the placement of the SHP gene at a new position on the genome of SPCSV relative to other closteroviridae. Northern analyses showed the presence of several sub-genomic RNAs, of which the accumulation was temporally regulated in infected tissues. The 5’-ends of seven sub-genomic RNAs were determined using a PCR based method, which indicated that the sgRNAs were capped

Horizontal gene transfer contributes to plant evolution : the case of Agrobacterium T-DNAs

Horizontal gene transfer (HGT) can be defined as the acquisition of genetic material from another organism without being its offspring. HGT is common in the microbial world including archaea and bacteria, where HGT mechanisms are widely understood and recognized as an important force in evolution. In eukaryotes, HGT now appears to occur more frequently than originally thought. Many studies are currently detecting novel HGT events among distinct lineages using next-generation sequencing. Most examples to date include gene transfers from bacterial donors to recipient organisms including fungi, plants, and animals. In plants, one well-studied example of HGT is the transfer of the tumor-inducing genes (T-DNAs) from some Agrobacterium species into their host plant genomes. Evidence of T-DNAs from Agrobacterium spp. into plant genomes, and their subsequent maintenance in the germline, has been reported in Nicotiana, Linaria and, more recently, in Ipomoea species. The transferred genes do not produce the usual disease phenotype, and appear to have a role in evolution of these plants. In this paper, we review previous reported cases of HGT from Agrobacterium, including the transfer of T-DNA regions from Agrobacterium spp. to the sweetpotato [ Ipomoea batatas (L.) Lam.] genome which is, to date, the sole documented example of a naturally-occurring incidence of HGT from Agrobacterium to a domesticated crop plant. We also discuss the possible evolutionary impact of T-DNA acquisition on plants

A Mixed Method Review and Comparative Analysis of Technology-Oriented Suicide Prevention Interventions for Adolescents and Adolescent Gatekeepers

This five-chapter dissertation compendium first introduces readers to the prevalence and impact of youth suicide, while also introducing readers to the important role gatekeepers can play in the prevention of youth suicide. Chapter one also describes the role of school personnel as gatekeepers who can be trained to address youth suicide prevention. Chapter two highlights technology-oriented suicide prevention interventions available to adolescents and adolescent gatekeepers. The efficacy and reach of these interventions, and the location and quality of supporting research evidence is explored and discussed. Chapter three describes federal and state guidelines regarding the training of school personnel as gatekeepers, and describes available evidence-based training programs included in the National Registry of Evidence-Based Programs and Practices. Given that there is variability in state-level policies and practices, five important gatekeeper training and policy considerations are discussed in relation to training duration, training frequency, training content, application of skills, and inclusion of technology. Chapter four further explores two technology-driven gatekeeper training programs in a side by side content analysis and head to head comparison with a sample of school personnel gatekeepers. Additional training considerations are also provided, with respect to online instructional quality, adult learning theory, social learning theory, behavioral rehearsal role-play practice, and strategic online classroom design, building upon the considerations outlined in chapter three. Chapter five addresses the importance of prioritizing technology-oriented gatekeeper training among school personnel, given the need for multifaceted solutions to youth suicide prevention. The dissertation compendium’s contribution to science and potential limitations and lessons learned are then discussed. Finally, the appendices outline supporting dissertation research study activities and other immediately relevant research documents. This dissertation compendium and dissertation research is timely, given the need for novel and scalable solutions to youth suicide prevention and present research gaps. The efficacy of technology-oriented adolescent and adolescent gatekeeper programs is outlined in chapter two, the efficacy of federal and state gatekeeper policies and training practices are outlined in chapter three, and chapter four further explores two online gatekeeper training programs in a school personnel sample while also connecting additional relevant training considerations. Together, these combined efforts permit exploration of important training and policy considerations across several levels. If school personnel are to assume their important role as gatekeepers, researchers have a duty to develop, rigorously evaluate, disseminate, and inform policies that maximize impact. Consequently, exploring program efficacy, federal and state policies, and school personnel training experiences and learning preferences is timely, relevant, and important. Data from this dissertation have potential to inform larger trials of training that may properly evaluate long-term effects on the population prevalence of youth suicidal behavior and contribute to training and policy level initiatives. The ultimate goal of this work is to ensure the dissemination and widespread use of high quality programs that effectively combat this critical public health problem

Anger in youth with anxiety:Relationship with reinforcement sensitivity, parental rejection and treatment outcome

Anxiety disorders are among the most prevalent disorders in children and young people (CYP). Around 20-30% of CYP with anxiety disorders also expresses oppositional behaviors. This subgroup presents with more severe mental health problems, worse psychosocial functioning, profits less from current treatments and has an increased risk of continued mental health problems later in life. In this dissertation, I investigated two pathways that may explain how CYP with anxiety disorders can develop behavioral problems. First, these comorbid behavioral problems may arise as a defensive response in for the CYP threatening situations, where they cannot avoid; especially for CYP with relatively high punishment sensitivity this may lead to behavioral problems. As a second possibility, behavioral problems could arise from frustration in situations where expected and desired rewards are not attained; especially for CYP with relative high reward sensitivity this may lead to behavioral problems. Additionally I investigated whether comorbid behavioral problems – via experiencing rejection from the environment – play a role in the maintenance of anxiety complaints. Lastly, it was investigated whether the treatment for anxiety would also reduce behavioral problems when they arise from experienced threat. For those where the behavioral problems arise out of frustration due to expected rewards that are not attained, it was expected that regular treatment for anxiety would not decrease the behavioral problems. This dissertation project provided consistent evidence for the role of frustration of not attaining expected rewards in comorbid behavioral problems. For the role of defensive responses herein, results were not univocal. In addition, the findings indicated that behavioral problems in adolescence indeed increased the risk of anxiety symptoms in young adulthood, although this appeared unrelated to experiencing more rejection from their parents. Importantly, behavioral problems generally decreased after anxiety-focused treatment, however not all CYP with behavioral problems had a reduction of their behavioral problems following treatment for anxiety. This seemed especially the case when the behavioral problems were not so much related to sensitivity for threat, but to the sensitivity for not attaining expected rewards. For CYP with this comorbid profile it seems important to add additional treatment modules focusing on the comorbid behavioral problems

Improving sweetpotato virus diagnostics

This flyer describes progress made between June 2014 and July 2015 to develop an easily accessible distribution map of all relevant viruses and virus strains infecting sweetpotato throughout SSA as well as appropriate diagnostic methods and protocols to detect them. It provides details about a generic virus detection method developed at CIP called small RNA sequencing and assembly (sRSA) to determine all viruses infecting sweetpotato in SSA: the pan-African sweetpotato Virome; as well as two diagnostic methods for detecting sweetpotato viruses: micro-arrays in a test tube (ClonDiag arrays) and an isothermal amplification method (LAMP)

THE HYDROSOCIAL COSTS OF HIGH-VOLUME HYDRAULIC FRACTURING: A TALE OF TWO COUNTIES IN MICHIGAN

Although natural gas has been praised as a clean and abundant energy source, the varying impacts and uncertainties surrounding the process of extracting natural gas from unconventional sources, known as horizontal high-volume hydraulic fracturing (HVHF) or “fracking,” have raised important concerns. The practice of HVHF is expanding so quickly that the full impacts are not yet known. This thesis project, using a grounded theory methodological approach, explores the risks and benefits associated with HVHF as recognized by the residents of two Michigan counties, one that currently produces natural gas by HVHF (Crawford County) and one that does not (Barry County). Through an analysis of media content related to HVHF in each case study site and interviews with stakeholders in both counties, this study examines perceptions of risks and benefits by comparing two communities that differ in their level of experience with HVHF operations, contributing to our understanding of how perceptions of risks and benefits are shaped by natural gas development. The comparative analysis of the case study counties revealed similarities and differences between the case study counties. Overall, Barry County residents identified fewer benefits and more risks, and had stronger negative perceptions than Crawford County residents. This study contributes to the social science literature by developing a richer theoretical frame for understanding perceptions of HVHF and also shares recommendations for industry, organizations, regulators, and government leaders interested in effectively communicating with community stakeholders about the benefits and risks of HVHF

The horizontal gene transfer of Agrobacterium T-DNAs into the series Batatas (genus Ipomoea) genome is not confined to hexaploid sweetpotato

The discovery of the insertion of IbT-DNA1 and IbT-DNA2 into the cultivated (hexaploid) sweetpotato [Ipomoea batatas (L.) Lam.] genome constitutes a clear example of an ancient event of Horizontal Gene Transfer (HGT). However, it remains unknown whether the acquisition of both IbT-DNAs by the cultivated sweetpotato occurred before or after its speciation. Therefore, this study aims to evaluate the presence of IbT-DNAs in the genomes of sweetpotato's wild relatives belonging to the taxonomic group series Batatas. Both IbT-DNA1 and IbT-DNA2 were found in tetraploid I. batatas (L.) Lam. and had highly similar sequences and at the same locus to those found in the cultivated sweetpotato. Moreover, IbT-DNA1 was also found in I. cordatotriloba and I. tenuissima while IbT-DNA2 was detected in I. trifida. This demonstrates that genome integrated IbT-DNAs are not restricted to the cultivated sweetpotato but are also present in tetraploid I. batatas and other related species

Diversidad fenotipica y genetica de cepas de Ralstonia solanacearum del Peru.

Ralstonia solacearum es el agente causal de la marchiez bacteriana. Esta enfermedad afecta numerosos cultivos de importancia economica especialmente en las regiones tropicales y sub tropicales. La amplia gama de hospsedantes, su distribucion y gran variabilidad del patogeno, hacen dificil el control de la enfermedad. Estudios sobre la variabilidad son importantes en mejoramiento para resistencia, competencia ecologica, manejo de la enfermedad y relaciones evolutivas y filogeneticas. El objetivo de este trabajo fue caracterizar fenotipica y genotipicamente 132 cepas de R. solanacearum aisladas de papa de distintos ecosistemas entre 1974 y 2013. La determinacion de biovares (Bv) se realizo en base a pruebas bioqumicas y se asigno el filotipo y secuevar correspondientemente mediante multiplex-PCR y- analisis de las consecuencias del gen endogluconasa. El analisis de datos se realizo por comparacion con las secuencias de las cepas de referencia depositadas en la base de datos del Gene Bank. Los resultados indican que el 100 % de las cepas del Peru corresponden al Filotipo II, el cual segun la clasificacion actual agrupa a las cepas originarias de America. Todas las cepas pertenecientes al biovar 2A (67.5 %) corresponden a los secuevares 1, 2. Este tipo de cepas tradicionamente presenta una baja variabilidad genetica y son las que afectan al cultivo de papa en zonas frias y templadas. El resto de las cepas analizadas fueron clasificadas dentro de los biovares (24.2%) y 2T (8.3%). Esta cepas presentaron una mayor variabilidad a nivel de secuencia y a la mayoria no se dudo asignar ningun secuevar conocido. Los resultados demuestan la diversidad de la poblacion R. solanacearum en el Peru. Infomacion valiosa que podria ayudar en las estrategias del control de la enfermedad