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- Creators: Arizona State University
- Creators: Chen, Qiang
- Member of: Theses and Dissertations
- Resource Type: Text
Description
ABSTRACT In terms of prevalence, human suffering and costs dengue infections are the most important arthropod-borne viral disease worldwide. Dengue virus (DENV) is a mosquito-borne flavivirus and the etiological agent of dengue fever and dengue hemorrhagic fever. Thus, development of a safe and efficient vaccine constitutes an urgent necessity. Besides the traditional strategies aim at generating immunization options, the usage of viral vectors to deliver antigenic stimulus in order to elicit protection are particularly attractive for the endeavor of a dengue vaccine. The viral vector (MVvac2) is genetically equivalent to the currently used measles vaccine strain Moraten, which adds practicality to my approach. The goal of the present study was to generate a recombinant measles virus expressing structural antigens from two strains of DENV (DENV2 and DENV4) The recombinant vectors replication profile was comparable to that of the parental strain and expresses either membrane bound or soluble forms of DENV2 and DENV4 E glycoproteins. I discuss future experiments in order to demonstrate its immunogenicity in our measles-susceptible mouse model.
ContributorsAbdelgalel, Rowida (Author) / Reyes del Valle, Jorge (Thesis advisor) / Hogue, Brenda (Committee member) / Frasch, Wayne D (Committee member) / Arizona State University (Publisher)
Created2013
Description
The concept of vaccination dates back further than Edward Jenner's first vaccine using cowpox pustules to confer immunity against smallpox in 1796. Nevertheless, it was Jenner's success that gave vaccines their name and made vaccinia virus (VACV) of particular interest. More than 200 years later there is still the need to understand vaccination from vaccine design to prediction of vaccine efficacy using mathematical models. Post-exposure vaccination with VACV has been suggested to be effective if administered within four days of smallpox exposure although this has not been definitively studied in humans. The first and second chapters analyze post-exposure prophylaxis of VACV in an animal model using v50ΔB13RMγ, a recombinant VACV expressing murine interferon gamma (IFN-γ) also known as type II IFN. While untreated animals infected with wild type VACV die by 10 days post-infection (dpi), animals treated with v50ΔB13RMγ 1 dpi had decreased morbidity and 100% survival. Despite these differences, the viral load was similar in both groups suggesting that v50ΔB13RMγ acts as an immunoregulator rather than as an antiviral. One of the main characteristics of VACV is its resistance to type I IFN, an effect primarily mediated by the E3L protein, which has a Z-DNA binding domain and a double-stranded RNA (dsRNA) binding domain. In the third chapter a VACV that independently expresses both domains of E3L was engineered and compared to wild type in cells in culture. The dual expression virus was unable to replicate in the JC murine cell line where both domains are needed together for replication. Moreover, phosphorylation of the dsRNA dependent protein kinase (PKR) was observed at late times post-infection which indicates that both domains need to be linked together in order to block the IFN response. Because smallpox has already been eradicated, the utility of mathematical modeling as a tool for predicting disease spread and vaccine efficacy was explored in the last chapter using dengue as a disease model. Current modeling approaches were reviewed and the 2000-2001 dengue outbreak in a Peruvian region was analyzed. This last section highlights the importance of interdisciplinary collaboration and how it benefits research on infectious diseases.
ContributorsHolechek, Susan A (Author) / Jacobs, Bertram L (Thesis advisor) / Castillo-Chavez, Carlos (Committee member) / Frasch, Wayne (Committee member) / Hogue, Brenda (Committee member) / Stout, Valerie (Committee member) / Arizona State University (Publisher)
Created2011
Biological and immunological characterization of plant-produced HIV-1 Gag/dgp41 virus-like particles
Description
Anti-retroviral drugs and AIDS prevention programs have helped to decrease the rate of new HIV-1 infections in some communities, however, a prophylactic vaccine is still needed to control the epidemic world-wide. Despite over two decades of research, a vaccine against HIV-1 remains elusive, although recent clinical trials have shown promising results. Recent successes have focused on highly conserved, mucosally-targeted antigens within HIV-1 such as the membrane proximal external region (MPER) of the envelope protein, gp41. MPER has been shown to play critical roles in the viral mucosal transmission, though this peptide is not immunogenic on its own. Gag is a structural protein configuring the enveloped virus particles, and has been suggested to constitute a target of the cellular immunity potentially controlling the viral load. It was hypothesized that HIV-1 enveloped virus-like particles (VLPs) consisting of Gag and a deconstructed form of gp41 comprising the MPER, transmembrane, and cytoplasmic domains (dgp41) could be expressed in plants. Plant-optimized HIV-1 genes were constructed and expressed in Nicotiana benthamiana by stable transformation, or transiently using a tobacco mosaic virus-based expression system or a combination of both. Results of biophysical, biochemical and electron microscopy characterization demonstrated that plant cells could support not only the formation of HIV-1 Gag VLPs, but also the accumulation of VLPs that incorporated dgp41. These particles were purified and utilized in mice immunization experiments. Prime-boost strategies combining systemic and mucosal priming with systemic boosting using two different vaccine candidates (VLPs and CTB-MPR - a fusion of MPER and the B-subunit of cholera toxin) were administered to BALB/c mice. Serum antibody responses against both the Gag and gp41 antigens could be elicited in mice systemically primed with VLPs and these responses could be recalled following systemic boosting with VLPs. In addition, mucosal priming with VLPs allowed for a robust boosting response against Gag and gp41 when boosted with either candidate. Functional assays of these antibodies are in progress to test the antibodies' effectiveness in neutralizing and preventing mucosal transmission of HIV-1. This immunogenicity of plant-based Gag/dgp41 VLPs represents an important milestone on the road towards a broadly-efficacious and inexpensive subunit vaccine against HIV-1.
ContributorsKessans, Sarah (Author) / Mor, Tsafrir S (Thesis advisor) / Matoba, Nobuyuki (Committee member) / Mason, Hugh (Committee member) / Hogue, Brenda (Committee member) / Fromme, Petra (Committee member) / Arizona State University (Publisher)
Created2011
Description
African Swine Fever (ASF), endemic in many African countries, is now spreading to other continents. Though ASF is capable of incurring serious economic losses in affected countries, no vaccine exists to provide immunity to animals. Disease control relies largely on rapid diagnosis and the implementation of movement restrictions and strict eradication programs. Developing a scalable, accurate and low cost diagnostic for ASF will be of great help for the current situation. CIM's 10K random peptide microarray is a new high-throughput platform that allows systematic investigations of immune responses associated with disease and shows promise as a diagnostic tool. In this study, this new technology was applied to characterize the immune responses of ASF virus (ASFV) infections and immunizations. Six sets of sera from ASFV antigen immunized pigs, 6 sera from infected pigs and 20 sera samples from unexposed pigs were tested and analyzed statistically. Results show that both ASFV antigen immunized pigs and ASFV viral infected pigs can be distinguished from unexposed pigs. Since it appears that immune responses to other viral infections are also distinguishable on this platform, it holds the potential of being useful in developing a new ASF diagnostic. The ability of this platform to identify specific ASFV antibody epitopes was also explored. A subtle motif was found to be shared among a set of peptides displaying the highest reactivity for an antigen specific antibody. However, this motif does not seem to match with any antibody epitopes predicted by a linear antibody epitope prediction.
ContributorsXiao, Liang (Author) / Sykes, Kathryn (Thesis advisor) / Zhao, Zhan-Gong (Committee member) / Stafford, Phillip (Committee member) / Arizona State University (Publisher)
Created2011
Description
Beliefs about change reflect how we understand phenomena and what kind of predictions we make for the future. Cyclical beliefs about change state that events are in a constant flux, and change is inevitable. Linear beliefs about change state that events happen in a non-fluctuating pattern and change is not commonplace. Cultural differences in beliefs about change have been documented across various domains, but research has yet to investigate how these differences may affect health status predictions. The present study addresses this gap by inducing different beliefs about change in a European-American college sample. Health status predictions were measured in terms of predicted likelihood of exposure to the flu virus, of contraction of the flu, and of receiving a flu vaccine. Most differences were observed among those who have a recent history of suffering from the flu. Among them, cyclical thinkers tended to rate their likelihood for exposure and contraction to be higher than linear thinkers. However, linear thinkers indicated that they were more likely to receive a flu vaccine. The different patterns suggest the possibility that cyclical beliefs may activate concepts related to cautionary behaviors or pessimistic biases, while linear beliefs may activate concepts related to taking action and exercising control over the environment. Future studies should examine the interplay between beliefs about change and the nature of the predicted outcome.
ContributorsKim, Summer Hyo Yeon (Author) / Kwan, Virginia S. Y. (Thesis advisor) / Neuberg, Steven L. (Committee member) / Cohen, Adam B. (Committee member) / Arizona State University (Publisher)
Created2012
Description
Plants are a promising upcoming platform for production of vaccine components and other desirable pharmaceutical proteins that can only, at present, be made in living systems. The unique soil microbe Agrobacterium tumefaciens can transfer DNA to plants very efficiently, essentially turning plants into factories capable of producing virtually any gene. While genetically modified bacteria have historically been used for producing useful biopharmaceuticals like human insulin, plants can assemble much more complicated proteins, like human antibodies, that bacterial systems cannot. As plants do not harbor human pathogens, they are also safer alternatives than animal cell cultures. Additionally, plants can be grown very cheaply, in massive quantities.
In my research, I have studied the genetic mechanisms that underlie gene expression, in order to improve plant-based biopharmaceutical production. To do this, inspiration was drawn from naturally-occurring gene regulatory mechanisms, especially those from plant viruses, which have evolved mechanisms to co-opt the plant cellular machinery to produce high levels of viral proteins. By testing, modifying, and combining genetic elements from diverse sources, an optimized expression system has been developed that allows very rapid production of vaccine components, monoclonal antibodies, and other biopharmaceuticals. To improve target gene expression while maintaining the health and function of the plants, I identified, studied, and modified 5’ untranslated regions, combined gene terminators, and a nuclear matrix attachment region. The replication mechanisms of a plant geminivirus were also studied, which lead to additional strategies to produce more toxic biopharmaceutical proteins. Finally, the mechanisms employed by a geminivirus to spread between cells were investigated. It was demonstrated that these movement mechanisms can be functionally transplanted into a separate genus of geminivirus, allowing modified virus-based gene expression vectors to be spread between neighboring plant cells. Additionally, my work helps shed light on the basic genetic mechanisms employed by all living organisms to control gene expression.
In my research, I have studied the genetic mechanisms that underlie gene expression, in order to improve plant-based biopharmaceutical production. To do this, inspiration was drawn from naturally-occurring gene regulatory mechanisms, especially those from plant viruses, which have evolved mechanisms to co-opt the plant cellular machinery to produce high levels of viral proteins. By testing, modifying, and combining genetic elements from diverse sources, an optimized expression system has been developed that allows very rapid production of vaccine components, monoclonal antibodies, and other biopharmaceuticals. To improve target gene expression while maintaining the health and function of the plants, I identified, studied, and modified 5’ untranslated regions, combined gene terminators, and a nuclear matrix attachment region. The replication mechanisms of a plant geminivirus were also studied, which lead to additional strategies to produce more toxic biopharmaceutical proteins. Finally, the mechanisms employed by a geminivirus to spread between cells were investigated. It was demonstrated that these movement mechanisms can be functionally transplanted into a separate genus of geminivirus, allowing modified virus-based gene expression vectors to be spread between neighboring plant cells. Additionally, my work helps shed light on the basic genetic mechanisms employed by all living organisms to control gene expression.
ContributorsDiamos, Andy (Author) / Mason, Hugh S (Thesis advisor) / Mor, Tsafrir (Committee member) / Hogue, Brenda (Committee member) / Stout, Valerie (Committee member) / Arizona State University (Publisher)
Created2017
Description
Is it possible to treat the mouth as a natural environment, and determine new methods to keep the microbiome in check? The need for biodiversity in health may suggest that every species carries out a specific function that is required to maintain equilibrium and homeostasis within the oral cavity. Furthermore, the relationship between the microbiome and its host is mutually beneficial because the host is providing microbes with an environment in which they can flourish and, in turn, keep their host healthy. Reviewing examples of larger scale environmental shifts could provide a window by which scientists can make hypotheses. Certain medications and healthcare treatments have been proven to cause xerostomia. This disorder is characterized by a dry mouth, and known to be associated with a change in the composition, and reduction, of saliva. Two case studies performed by Bardow et al, and Leal et al, tested and studied the relationships of certain medications and confirmed their side effects on the salivary glands [2,3]. Their results confirmed a relationship between specific medicines, and the correlating complaints of xerostomia. In addition, Vissink et al conducted case studies that helped to further identify how radiotherapy causes hyposalivation of the salivary glands [4]. Specifically patients that have been diagnosed with oral cancer, and are treated by radiotherapy, have been diagnosed with xerostomia. As stated prior, studies have shown that patients having an ecologically balanced and diverse microbiome tend to have healthier mouths. The oral cavity is like any biome, consisting of commensalism within itself and mutualism with its host. Due to the decreased salivary output, caused by xerostomia, increased parasitic bacteria build up within the oral cavity thus causing dental disease. Every human body contains a personalized microbiome that is essential to maintaining health but capable of eliciting disease. The Human Oral Microbiomics Database (HOMD) is a set of reference 16S rRNA gene sequences. These are then used to define individual human oral taxa. By conducting metagenomic experiments at the molecular and cellular level, scientists can identify and label micro species that inhabit the mouth during parasitic outbreaks or a shifting of the microbiome. Because the HOMD is incomplete, so is our ability to cure, or prevent, oral disease. The purpose of the thesis is to research what is known about xerostomia and its effects on the complex microbiome of the oral cavity. It is important that researchers determine whether this particular perspective is worth considering. In addition, the goal is to create novel experiments for treatment and prevention of dental diseases.
ContributorsHalcomb, Michael Jordan (Author) / Chen, Qiang (Thesis director) / Steele, Kelly (Committee member) / Barrett, The Honors College (Contributor) / College of Letters and Sciences (Contributor)
Created2015-05
Description
Agrobacterium tumefaciens has the ability to transfer its tumor inducing (Ti) plasmid into plant cells. In the last decade, agroinfiltration of Nicotiana benthamiana plants has shown promising results for recombinant protein production. However, A. tumefaciens produce endotoxins in the form of lipopolysaccharides (LPS), a component of their outer membrane that can induce organ failure and septic shock. Therefore, we aimed to detoxify A. tumefaciens by modifying their Lipid A structure, the toxic region of LPS, via mutating the genes for lipid A biosynthesis. Two mutant strains of A. tumefaciens were infiltrated into N. benthamiana stems to test for tumor formation to ensure that the detoxifying process did not compromise the ability of gene transfer. Our results demonstrated that A. tumefaciens with both single and double mutations retained the ability to form tumors. Thus, these mutants can be utilized to generate engineered A. tumefaciens strains for the production of plant-based pharmaceuticals with low endotoxicity.
ContributorsHaseefa, Fathima (Author) / Chen, Qiang (Thesis director) / Mason, Hugh (Committee member) / Hurtado, Jonathan (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Vaccines against the arthropod-borne dengue virus (DENV) are still commercially nonexistent. A subunit immunization strategy may be of value, especially if a safe viral vector acts as a biologically active adjuvant. The DENV envelope protein (E), the main target for neutralizing immune responses, has three conformational domains. The immunoglobulin-like and independently folding domain III (DIII) contains epitopes that elicit highly specific neutralizing antibodies. The hepatitis B small surface antigen (HBsAg, S) was used as a scaffold to display DENV 2 DIII on a virus-like particle (VLP). A measles virus (MV) was engineered to vector HBsAg and the hybrid glycoprotein DIII-HBsAg in two different loci (DIII-S). Despite the relatively deleterious effect on replication caused by the insertion of two transcription cassettes, the recombinant virus MVvac2(DIII-S,S)P induced the secretion of DIII-S hybrid VLP with a similar sucrose density as HBsAg particles (1.10-1.12g/ml) and peaked at 48 h post-infection producing 1.3x106 TCID50/ml infectious MV units in vitro. A second recombinant virus, MVvac2(DIII-S)N, was engineered to vector only the hybrid DIII-S. However, it did not induce the secretion of hybrid HBsAg particles in the supernatant of infected cells. The immunogenicity of the recombinant viruses was tested in a MV-susceptible small animal model, the experimental group which received two 105 TCID50 I.P. doses of MVvac2(DIII-S,S)P in a 28 day interval developed a robust immune response against MV (1:1280), HBsAg (787 mIU/ml) and DENV2 (Log10 neutralization index of 1.2) on average. In summary, it is possible to display DENV E DIII on hybrid HBsAg particles vectored by MV that elicit an immune response. This forms the basis for a potential vaccine platform against DENV.
ContributorsHarahap, Indira (Author) / Reyes del Valle, Jorge (Thesis advisor) / Hogue, Brenda G (Thesis advisor) / Lake, Douglas (Committee member) / Mason, Hugh (Committee member) / Arizona State University (Publisher)
Created2015
Description
Background: This study examines how pro-vaccine flu messages, guided by the Extended Parallel Process Model (EPPM), affect parents’ intentions to vaccinate their children.
Methods: Parents of children six months to five years old (N = 975) were randomly exposed to one of four high-threat/high-efficacy messages (narrative, statistical, combined, control) and completed a follow-up survey. Differences between message conditions were assessed with one-way ANOVAs, and binary logistic regressions were used to show how constructs predicted intentions.
Results: There were no significant differences in the ANOVA results at p = .05 for EPPM variables or risk EPPM variables. There was a significant difference between message conditions for perceived manipulation (p = 0.026), authority, (p = 0.024), character (p = 0.037), attention (p < .000), and emotion (p < .000). The EPPM model and perceptions of message model (positively), and the risk EPPM model and fear control model (negatively), predicted intentions to vaccinate. Significant predictor variables in each model at p < .05 were severity (aOR = 1.83), response efficacy (aOR = 4.33), risk susceptibility (aOR = 0.53), risk fear (aOR = 0.74), issue derogation (aOR = 0.63), perceived manipulation (aOR = 0.64), character (aOR = 2.00), and personal relevance (aOR = 1.88). In a multivariate model of the significant predictors, only response efficacy significantly predicted intentions to vaccinate (aOR = 3.43). Compared to the control, none of the experimental messages significantly predicted intentions to vaccinate. The narrative and combined conditions significantly predicted intentions to search online (aOR = 2.37), and the combined condition significantly predicted intentions to talk to family/friends (aOR = 2.66).
Conclusions: The EPPM may not be effective in context of a two-way threat. Additional constructs that may be useful in the EPPM model are perceptions of the message and fear control variables. One-shot flu vaccine messages will be unlikely to directly influence vaccination rates; however they may increase information-seeking behavior. The impact of seeking more information on vaccination uptake requires further research. Flu vaccine messages should be presented in combined form. Future studies should focus on strategies to increase perceptions of the effectiveness of the flu vaccine.
Methods: Parents of children six months to five years old (N = 975) were randomly exposed to one of four high-threat/high-efficacy messages (narrative, statistical, combined, control) and completed a follow-up survey. Differences between message conditions were assessed with one-way ANOVAs, and binary logistic regressions were used to show how constructs predicted intentions.
Results: There were no significant differences in the ANOVA results at p = .05 for EPPM variables or risk EPPM variables. There was a significant difference between message conditions for perceived manipulation (p = 0.026), authority, (p = 0.024), character (p = 0.037), attention (p < .000), and emotion (p < .000). The EPPM model and perceptions of message model (positively), and the risk EPPM model and fear control model (negatively), predicted intentions to vaccinate. Significant predictor variables in each model at p < .05 were severity (aOR = 1.83), response efficacy (aOR = 4.33), risk susceptibility (aOR = 0.53), risk fear (aOR = 0.74), issue derogation (aOR = 0.63), perceived manipulation (aOR = 0.64), character (aOR = 2.00), and personal relevance (aOR = 1.88). In a multivariate model of the significant predictors, only response efficacy significantly predicted intentions to vaccinate (aOR = 3.43). Compared to the control, none of the experimental messages significantly predicted intentions to vaccinate. The narrative and combined conditions significantly predicted intentions to search online (aOR = 2.37), and the combined condition significantly predicted intentions to talk to family/friends (aOR = 2.66).
Conclusions: The EPPM may not be effective in context of a two-way threat. Additional constructs that may be useful in the EPPM model are perceptions of the message and fear control variables. One-shot flu vaccine messages will be unlikely to directly influence vaccination rates; however they may increase information-seeking behavior. The impact of seeking more information on vaccination uptake requires further research. Flu vaccine messages should be presented in combined form. Future studies should focus on strategies to increase perceptions of the effectiveness of the flu vaccine.
ContributorsHall, Sarah (Author) / Jehn, Megan (Thesis advisor) / Mongeau, Paul (Committee member) / Hruschka, Daniel (Committee member) / Margolis, Eric (Committee member) / Arizona State University (Publisher)
Created2015