Theses and Dissertations

Permanent URI for this collection

https://hdl.handle.net/10217/100474

Browse

Now showing 1 - 6 of 6

Open Access
Computational tools to identify correlates of vaccine-induced protection against tuberculosis
(Colorado State University. Libraries, 2021) Fox, Amy, author; Henao-Tamayo, Marcela, advisor; Anderson, Brooke, advisor; Abdo, Zaid, committee member; Fosdick, Bailey, committee member
Tuberculosis is a significant threat to human health. While the BCG vaccine exists to protect children from disseminated forms of tuberculosis, it fails to protect against pulmonary tuberculosis. Thus, a better vaccine is needed. However, the immune system in response to tuberculosis and the BCG vaccine is incompletely understood. We sought to develop novel analysis methods to help understand the immune system. This dissertation describes an analysis tool, cyto-feature engineering, that rapidly identifies flow cytometry immune cell populations utilizing experimental controls. The tool was corroborated through testing the pipeline on different types of flow cytometry datasets. Cyto-feature engineering was then utilized to understand the immune response to two immunomodulatory drugs—losartan and propranolol—when used in conjunction with the BCG vaccine. This study identified an increase in T cell responses due to drug administration, but ultimately failed to decrease bacterial burden in the lung and spleen. Other studies employed a new method for identifying immune cells correlated with various metabolites in the context of tuberculosis. The method can be utilized to generate hypotheses from secondary data sources and gain new biological insight. Using this method, we identified a potential correlation between CD45RA and arachidonic acid metabolism which could serve as a potential target for future vaccination studies. The research outlined in this dissertation will hopefully lead to better immunological analyses of data and the development of a better tuberculosis vaccine.
Open Access
Development of Lactobacillus acidophilus as an oral vaccine vector and effects of rice bran ingestion on the mucosal health of Malian infants
(Colorado State University. Libraries, 2020) Vilander, Allison C., author; Dean, Gregg, advisor; Abdo, Zaid, committee member; Dow, Steven, committee member; MacNeill, Amy, committee member; Ryan, Elizabeth, committee member
Most pathogens enter the body at the mucosa and induce innate and adaptive immune responses at these surfaces essential for protection against infection and disease. Induction of mucosal immune responses is best achieved locally but mucosal vaccines have been difficult to develop with few currently approved for use. Almost all are attenuated live vaccines which limits their use and efficacy in some populations. Strategies to enhance the mucosal immune response to vaccination and move away from attenuated live vaccines are needed. Prebiotics (nondigestible food ingredients that promotes growth of beneficial microorganisms) and probiotics (live microorganisms that are beneficial when ingested) are an active area of interest for improving mucosal health and increasing oral vaccine performance. Here we present the development of the probiotic Gram-positive lactic acid bacteria Lactobacillus acidophilus (LA) as a novel oral subunit vaccine. LA has many advantages as an oral vaccine vector including endogenous acid and bile resistance, heat tolerance, and numerous proteins that interact with the mucosal immune system. We show that LA can induce immune responses to weakly immunogenic neutralizing peptides from HIV-1 and rotavirus. To enhance the immune response, we developed the E. coli type I pilus protein, FimH, as a LA vaccine adjuvant. FimH increased the immune response to vaccination and increased LA trafficking by antigen presenting cells to the mesenteric lymph node, an important site of mucosal immune induction. We also evaluate the effects of ingestion of the nutrient dense prebiotic rice bran on mucosal health in a cohort of healthy Malian infants at risk for malnutrition and the subclinical condition environmental enteric dysfunction. Rice bran ingestion was found to decrease episodes of diarrhea, decrease the age to elevated fecal microbiome α-diversity, and stabilize total fecal secretory IgA concentrations over time. These results indicate that rice bran protects from diarrhea and improves the mucosal environment.
Open Access
Epidemiological investigation of antimicrobial resistance in beef production using metagenomic sequencing
(Colorado State University. Libraries, 2019) Doster, Enrique, author; Hoover, Edward A., advisor; Morley, Paul S., advisor; Belk, Keith E., committee member; Abdo, Zaid, committee member; Gow, Sheryl P., committee member
Globally, the emergence of antimicrobial resistance (AMR) resulting in treatment failure is recognized as a growing public health threat. Antimicrobial use practices used in beef production are thought to be a direct driver of increasing antimicrobial resistance in pathogens and the environment, in part due to the higher volumes of antimicrobial drug necessary to treat cattle weighing 10 times more than an average person. This has led policy makers and public health organizations to promote "judicious use" or outright ban of antimicrobial drugs in livestock production. Use of antimicrobials is unavoidable for the treatment of disease and we must therefore learn how we can best adjust our AMD use to reduce selection of AMR pathogens. However, outside of important indicator organisms and pathogens, little is known about how different antimicrobial drug use practices affect communities of microorganisms, or microbiomes, and the AMR gene determinants, or resistome, shared between pathogen and non-pathogens alike. With advances in high-throughput sequencing (HTS), we can perform culture-independent studies and gain a better understanding of how antimicrobial drug use practices in livestock production affect AMR epidemiology. This dissertation consists of five studies that employ HTS to characterize the microbiome and resistome of samples with differing AMD exposure along the beef production line. Projects begin with a look into the short-term effects on the microbiome and resistome of feedlot cattle following treatment with a macrolide drug, tulathromycin, in the manuscript "Investigating Effects of Tulathromycin Metaphylaxis on the Fecal Resistome and Microbiome of Commercial Feedlot Cattle Early in the Feeding Period". Fecal samples collected in this project also were processed with aerobic culture, polymerase chain reaction (PCR), and lateral flow immunoassay for identification of Salmonella enterica and the comparison of these results are presented in "A Cautionary Report for Pathogen Identification Using Shotgun Metagenomics; a Comparison to Aerobic Culture and Polymerase Chain Reaction for Salmonella enterica Identification". Samples collected as part of a longitudinal study in feedlot cattle were analyzed to characterize the associations between AMD use and AMR in two bacterial species. These archived samples are leveraged for a broader understanding of AMR dynamics by adding a community-level perspective to results from aerobic culture. Results in individual cattle are presented in "Antimicrobial Drug Use in Beef Feedlots; Effects on the Microbiome and Resistome Dynamics in Individual Cattle" and results at the pen-level in "Metagenomic Investigation of the Effects of Antimicrobial Drug Use Practices on the Microbiome and Resistome of Beef Feedlot Cattle". Finally, in "Metagenomic Characterization of the Microbiome and Resistome in Retail Ground Beef" we examined the end of the beef production line by comparing the microbiome and resistome of retail ground beef products from either conventional production systems or those labeled as "raised with antibiotics" (RWA). The five studies presented in this dissertation each contribute to the collective understanding of how AMD use in livestock production system can affect the ecology of AMR in microbial communities. These projects are useful first steps in learning to manage AMR in beef production systems; encompassing a targeted look at the use of one type of AMD, characterizing the resistome dynamics in individual cattle and pens over time in a feedlot, a comparison of the resistome in ground beef products, and many other aspects of AMR epidemiology. The final study, describing limits to incorporating HTS for pathogen identification, serves as a cautionary reminder that with new technologies come new challenges and that research must keep pace.
Open Access
From BCG vaccination routes to lung and gut microbiota: avenues to tackle Mycobacterium tuberculosis infection
(Colorado State University. Libraries, 2021) Silva-Angulo, Fabiola, author; Henao-Tamayo, Marcela, advisor; Weir, Tiffany, committee member; Abdo, Zaid, committee member; Izzo, Angelo, committee member
Tuberculosis is an infectious lung disease responsible for approximately 1.4 million human deaths, world-wide every year. The causal agent of tuberculosis, Mycobacterium tuberculosis (M. tuberculosis), has been estimated to latently infect one-third of the human population. Currently, the BCG vaccine, a live attenuated strain of Mycobacterium bovis, is the only vaccine available to control the disease. Although the BCG vaccine has been the most widely administered worldwide and has been used for more than 100 years, tuberculosis dissemination remains uncontrolled and highly prevalent, especially in developing countries. Several questions about the effect that local microbiota and the administration route of BCG vaccination make on tuberculosis immunopathogenesis remain unanswered. These questions are critical for developing new approaches to control the disease. BCG vaccination is administered intradermally, however, some studies have suggested that BCG vaccination efficacy may be dependent on the administration route. Vaccination through the natural route of M. tuberculosis infection and a combination of other routes have been studied in animal models with varying results. Currently, the analysis of vaccination through the natural infection site is an attractive approach to priming innate immunity. The first study of this thesis examined the immune response induced after BCG vaccination using different routes (aerosol, subcutaneous, intravenous, and intranasal) in C57BL/6 mic and their response to pulmonary M. tuberculosis infection. The study was focused on specific markers of both CD4+ and CD8+ T cells. Our data suggested differences in the adaptive immune response based on the route of BCG vaccination and mainly elicited by CD4+ T cell immune response, with the intranasal delivery the most effective in decreasing the growth of M. tuberculosis in lungs. Another crucial question is the effect of M. tuberculosis infection and BCG vaccination on the structure, diversity, and potential function of the host lung and gut microbiota. Thus, the objective for the second study of this thesis was to characterize the effect of BCG vaccination and M. tuberculosis infection on the lung and gut micro- and mycobiota of C57BL/6 mice. Results indicated that BCG vaccination and M. tuberculosis infection in mice altered the relative lung abundance of Firmicutes and Bacteroidetes phyla compared to the control non-vaccinated, non-infected group. Lung diversity was most affected after M. tuberculosis infection. A multivariate regression approach was used to compare the profile evolution of gut and lung microbiota. More genera had modified relative abundances associated with BCG vaccination status at the gut level compared with lung. Conversely, genera with modified relative abundances associated with M. tuberculosis infection were numerous at lung level, and indicated that the local host response against infection impacted the whole microbial flora while the immune response after vaccination modified mainly the gut microbiota. This study demonstrated that parenteral vaccination with a live attenuated microorganism induced both lung and gut dysbiosis, which may play a crucial role in the immune response to M. tuberculosis infection.
Open Access
Investigation of RelBE1 toxin-antitoxin function in the carbon-dependent metabolic adaptation of Mycobacterium tuberculosis
(Colorado State University. Libraries, 2022) Starkey, Julie M., author; Slayden, Richard, advisor; Dobos, Karen, committee member; Abdo, Zaid, committee member; Tjalkens, Ron, committee member
Tuberculosis (TB) is a devastating disease with suboptimal treatment regimens and a single vaccine with variable efficacy. Reducing the global burden of TB requires a refined arsenal of methods to prevent and treat the disease, which necessitates a better understanding of M. tuberculosis (Mtb) pathogenesis during infection. Mtb undergoes continuous metabolic reprogramming throughout acute and chronic stages of infection in order to survive and persist harsh host conditions, and the regulatory network responsible for mediating metabolic adaptation has not been fully defined. Mtb harbors at least 88 Toxin-antitoxin (TA) loci that have been proposed to function as regulatory modules in response to stress. TA systems are uniquely abundant in Mtb, making them viable targets for the treatment of both active and latent infection. Several RelBE TA systems are present in Mtb, and the RelE toxins function as ribonucleases to inhibit translation when not bound to RelB antitoxins. The genes encoding relBE1 are adjacent to a gene that encodes an enzyme involved in central carbon metabolism, which could suggest a regulatory role for RelBE1 in carbon metabolism. We aimed to explore the relationship between the RelBE1 TA system and carbon-mediated metabolic adaptation. This work incorporated in vitro transcriptional and genetic studies under defined carbon sources to investigate the activity of RelBE1 and the requirement of RelE1 in Mtb metabolism, growth, and viability in the presence of different carbon sources. We observed transcriptional and physiological trends consistent with the hypothesis that RelBE1 contributes to iii adaptation of Mtb metabolism in the presence of cholesterol and oleate. Additionally, we found evidence that supports the necessity of RelE1 in Mtb metabolism under conditions depleted of nutrients. To investigate if multiple RelBE systems work redundantly or cooperatively in Mtb metabolic adaptation, we applied CRISPRi to simultaneously silence three RelBE TA loci. CRISPRi construction of knockdown mutants resulted in variable success but did not fully resolve the question regarding the cooperative or redundant functions of RelBE systems in Mtb metabolism. Nonetheless, the study provided the building blocks for efficient genetic manipulation of multiple TA systems in Mtb that are essential for exploring the coordination of TA systems in their contribution to Mtb pathogenesis. This thesis work contributes to the debate regarding TA system function in Mtb stress response and adaptation during infection. Given the limitations of the presented studies, further work is warranted to elucidate the relationship between TA systems and Mtb pathogenesis. Expanding our understanding of TA systems in TB disease would provide novel avenues in research to improve treatments against TB.
Embargo
The adventures of Lactobacillus acidophilus: evaluating a recombinant probiotic rotavirus vaccine from host and microbial perspectives
(Colorado State University. Libraries, 2024) Gilfillan, Darby L., author; Vilander, Allison, advisor; Dean, Gregg, advisor; Abdo, Zaid, committee member; Wilkins, Mike, committee member
Rotavirus is an enteric infection of global importance causing diarrheal-associated illness that can be fatal in young children and the elderly. There is a gap in vaccine efficacy between high- and lower-middle-income countries (LMIC) with LMIC often experiencing diminished vaccine-conferred protection. Rotaviruses, whether attenuated vaccine strains or primary pathogens, do not exist in isolation within the host's gastrointestinal tract. Other actors present within the microbiome can inhibit or augment vaccine efficacy by influencing the vaccine itself or the mucosal immune response. Understanding and exploiting interactions between host and microbe is a promising frontier for mucosal vaccinology. This dissertation will explore the probiotic Lactobacillus acidophilus (LA) as a vaccine platform for a microbiome-minded, next-generation approach to rotavirus immunization. We developed and confirmed a novel recombinant LA (rLA) vaccine expressing rotavirus antigens of the VP8* domain from the rotavirus EDIM VP4 capsid protein along with the adjuvants FimH and FliC. Rotavirus naïve adult BALB/cJ mice were orally immunized followed by murine rotavirus strain ECWT viral challenge. Antirotavirus serum IgG and antigen-specific antibody-secreting cell responses were detected in rLA-vaccinated mice. A day after the oral rotavirus challenge, fecal antigen shedding was significantly decreased in the rLA group. These results demonstrate the potential of rLA platforms to generate protective mucosal immunity. Additionally, metagenomic and metatranscriptomic analyses of exogenous probiotic administration within the murine small intestine revealed differences between LA genome expression and the whole metatranscriptome in recombinant- versus wild-type LA-vaccinated mice. LA genome expression in rLA-vaccinated mice had decreased carbohydrate metabolism and increased stress responses. We also detected antigen and adjuvant transcript expression only in mice exposed to the rLA platform. There was relative enrichment of probiotic species in the wild-type group with overall increased α- and β-diversity in the buffer compared to probiotic groups. These results highlight the interactions between an exogenous probiotic and the host microbiome at an immune inductive site. Finally, we used an in vitro model to evaluate modulation of polyunsaturated fatty acid (PUFA) metabolism on host cell and (r)LA interactions. Both (r)LA and PUFA treatments significantly changed pathogen recognition receptor expression. (r)LA treatment mainly altered inflammatory cytokine expression while PUFA supplementation primarily influenced mucin expression. rLA strains adhered more to host cells than wild-type LA while the rLA strain expressing both antigens and adjuvants may better prevent E. coli adhesion. These results and methodologies provide a starting point for further investigation into PUFA metabolism as a mechanism for improving rLA immunogenicity and competition against other enteric pathogens.

Browse

Browsing Theses and Dissertations by Author "Abdo, Zaid, committee member"

Results Per Page

Sort Options