Theses and Dissertations at Montana State University (MSU)
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Item An interrogation of herpes simplex virus type-1 gene expression during neuronal infection(Montana State University - Bozeman, College of Agriculture, 2024) Domanico, Luke Frank; Chairperson, Graduate Committee: Matt Taylor; This is a manuscript style paper that includes co-authored chapters.Herpes Simplex virus-type-1 (HSV-1) is a ubiquitous human pathogen casually referred to as "the gift that keeps on giving". The seemingly benign recurring herpetic lesions caused by acute HSV-1 infection are an obnoxious reminder of an incurable infection. HSV-1 maintains lifelong persistence in the infected host through a unique form of infection in peripheral neurons, conventionally termed latency. The latently infected neuron acts as a viral reservoir and is the focal point of herpetic disease. The latent HSV-1 infection represents a brilliant orchestration of viral gene regulation, manipulation of highly polarized cells, and seamless evasion of immunological clearance. Though, the viral mechanisms and cellular factors that govern the establishment, maintenance and reactivation from latency are elusive and challenging to study. The work included here aims to uncover the cryptic factors involved in and supporting the latent HSV-1 infection. Authored publications include the demonstration of a recombinant HSV-1 that enables temporal discretion of viral gene expression, and the revelation of a stunning, yet obscure phenotype of neuronal infection. Next is the implementation of a single-cell culturing method using drop-based microfluidic technology to resolve HSV-1 infection in isolated neurons. Together, this work reveals that the early events of neuronal infection are critical to determining the lytic or latent outcome of infection. Inoculating dose impacts the kinetics of viral replication, and the establishment of lytic or latent HSV-1 infection. Furthermore, evaluation of viral gene expression during latent HSV-1 infection suggests that the distinction between lytic and latent HSV-1 infection is less mutually exclusive than is historically appreciated. Finally, I present preliminary and ongoing research suggesting that a cellular transcription factor called nuclear factor-kappa B (NF-kB) differentially engages in HSV-1 infection. NF-kB supports efficient lytic gene transcription in epithelial cells, while promoting the establishment of latent HSV-1 infection of neurons.Item Certification of reprocessing standardization: preventing endoscopy associated infections(Montana State University - Bozeman, College of Nursing, 2023) Ranck, Aaron Thomas; Chairperson, Graduate Committee: Yoshiko Yamashita ColcloughBackground: Between 2012-2015, 25 outbreaks were linked to contaminated duodenoscopes worldwide due to human error or negligence during reprocessing. A standardized education and training reprocessing program is necessary to address nationally increasing Endoscopy Associated Infections (EAIs). Problem: A Montana endoscopy unit committed to adopting national standards. An unmet objective was the certification of endoscopy reprocessing (CER) requirement. This pilot project sted in developing a mandatory CER protocol to optimize infection control. Methods: A Plan-Do-Study-Act (PDSA) quality improvement method was employed, with pre- and post-intervention design to evaluate infection incidence and risk. This involved patient chart review and duodenoscope reprocessing log review. An aggregate comparison was made between a CER participant and eight non-CER technicians. To inform future program development, a survey was conducted to collect data on CER participant motivation, preparation, and perception of recommended educational materials. Intervention: A pilot study involving a single participant attempting to obtain nationally recognized CER was conducted. The facility manager shared the facility's intention of mandatory certification and via endoscope technician meeting, including incentives. Created certification-benefit video presentation and exam preparation materials were provided one month before the exam. Results: Zero infections occurred during pre- and post-intervention. Pre-intervention, the average infection risk scores per endoscope reprocessing were 12.2 relative light units (RLUs) for the unit and 11.1 RLUs for the CER participant. Post-intervention phase, these numbers were 15.7 RLUs by non-CER technicians and 2.8 RLUs by the CER participant. Conclusion: This study highlights the effectiveness of mandatory CER in reducing the risk of patient infections. Maintaining infection prevention and control in endoscopy procedures requires ongoing education, training, and motivation. Informing employees of the benefits of standardization may improve motivation.Item Virus-like particle surface toll-like receptor signaling modulates host response to bacterial infection(Montana State University - Bozeman, College of Agriculture, 2022) Hatton, Alexis Alexandria; Chairperson, Graduate Committee: Mark T. Quinn; This is a manuscript style paper that includes co-authored chapters.Innate immune recognition of viruses is critical for the rapid response and subsequent clearance of an infection. The primary focus of virus innate immune recognition is the recognition of viral nucleic acids post-infection. However, innate pattern recognition receptors (PRRs) have been demonstrated to recognize viral proteins independent of infection. One group of PRRs associated with viral recognition are Toll-like Receptors (TLRs). With the discovery of TLRs in the late 1990's, over two decades of research have endeavored to identify if cell surface TLRs recognize viral proteins and if so, understand whether surface TLR-viral recognition benefits the host or the virus. To the benefit of the host, it was previously determined that host-recognition of viral proteins protects mice from bacterial infection early post-virus exposure, independent of viral nucleic acids (virus-like particle; VLP). This suggested that early viral protein recognition could protectively prime the host against bacterial infection. Our investigation here attempts to address the generality of surface TLR-virus recognition independent of infection, how viral protein recognition alters the subsequent signaling response to bacterial infection, and finally, if/how expression system-associated variables interfere with the interpretation of our study. We utilized macrophage deficient in surface TLRs and TLR-associated signaling proteins to address the TLR signaling pathway responsible for the general response to VLPs that results in reduced bacterial burden. We found that different surface TLRs were responsible for reducing bacterial burden, resulting in the activation of different signaling pathways dependent upon the VLP macrophages were exposed to. In addition, our results demonstrate how expression system-associated variables alter the interpretation of signaling pathways activated by surface TLRs.Item Influenza D viruses in polymicrobial infections(Montana State University - Bozeman, College of Agriculture, 2022) Robinson, Emily Faith; Chairperson, Graduate Committee: Agnieszka Rynda-AppleInfluenza D viruses (IDVs) comprise Deltainfluenzaviridae, the newest genus of the Orthomyxoviridae family. This group of viruses primarily infects the upper respiratory tract and causes only mild symptoms. Unlike most influenza viruses, IDVs infect a wide range of ungulates, including cattle, swine, and sheep. In cattle, IDVs are thought to act as an etiologic agent in bovine respiratory disease (BRD), a severe polymicrobial disease primarily impacting calves. In chapters 2-4 of this thesis, we report the first experimental infection of an IDV in sheep with and without a recent Mycoplasma ovipneumoniae (M. ovipneumoniae) bacterial infection. We found no evidence of overt illness in IDV-infected lambs. Our findings suggested that recent M. ovipneumoniae infection induced a mild proinflammatory innate immune response that contributed to an enhanced neutralizing antibody response compared to that of M. ovipneumoniae-naive IDV-infected lambs. These findings suggest that although these lambs did not present with clinical symptoms in response to IDV, carriage could contribute to the inflammatory response in sheep experiencing polymicrobial infections. Influenza infections can confer short-term protection against additional viral pathogens. This process, called heterologous viral interference, is mediated by type I interferon antiviral signaling. This phenomenon predominantly occurs when a host is infected with a mild virus followed by a more severe virus. In humans, heterologous viral interference can result in an attenuation or delay of symptoms associated with the more severe pathogen. Evidence suggests that IDVs infect humans, although there is no evidence that these infections are symptomatic. To gain insight into whether IDVs can reduce the symptoms of influenza A virus (IAV), we performed a series of experiments in mice. In Chapter 5 of this thesis we show that IDV infection can, but does not always, reduce disease associated with IAV. We subsequently sought to identify critical type I interferon signaling events underlaying this phenomenon, but our results remain inconclusive.Item Evaluation of conditions and mechanisms of alphaherpesvirus superinfection exclusion(Montana State University - Bozeman, College of Agriculture, 2022) Cwick, James Patrick; Chairperson, Graduate Committee: Matt Taylor; This is a manuscript style paper that includes co-authored chapters.Superinfection exclusion (SIE) is a natural phenomenon where one virus prevents subsequent entry of another virus. Studies have shown its impact on viral replication and reproduction of many different viruses, where exclusion targets specific parts of the virus' lifecycle. Alphaherpesviruses, specifically pseudorabies virus (PRV) and Herpes Simplex virus type 1 (HSV-1), have demonstrated at least two forms of SIE. However, the form of SIE that occurs early in viral replication remains of interest due to both its timing and unknown mechanisms of regulation. Research on this topic will stimulate development of vaccines in global health by providing targets for disruption of alphaherpesvirus infection and information on unknown aspects of alphaherpesvirus infection. We have developed two models for assessing early SIE for alphaherpesviruses: a fluorescent reporter model to quantify virions within the cell. Both models contributed to data that determined our early SIE is influenced by multiplicity of infection (MOI) for both the primary and secondary virions. Imaging flow cytometry was utilized in conjunction with fluorescent microscopy as a possible means to study early SIE in large population samples. Subsequent data from my experiments indicates that cellular factors like cellular receptors, clathrin, and actin arrangement had minimal influence on early SIE of alphaherpesviruses. However, the results from capsid trafficking model in combination with experiments involving heparin and bortezomib indicate that early SIE differs in PRV and HSV-1. Data indicates that PRV SIE inhibits the step of viral entry/fusion, while HSV-1 SIE disrupts in a post-entry manner. Data obtained from this dissertation indicates that early SIE influences alphaherpesviruses differently and presents possible means to study antiviral techniques and methods.Item Investigating neutrophil cell fate following interactions with Staphylococcus aureus(Montana State University - Bozeman, College of Agriculture, 2022) Dankoff, Jennifer Grace; Chairperson, Graduate Committee: Jovanka Voyich-Kane; This is a manuscript style paper that includes co-authored chapters.Staphylococcus aureus is a ubiquitous pathogen with a growing list of antibiotic resistant capabilities. This gram-positive bacterium is able to cause a range of diseases, from a benign state of nasal colonization to fatal endocarditis. The ability to exist along this spectrum is largely dependent on the molecular dialog that takes place between the pathogen and the host, specifically white blood cells known as neutrophils. Neutrophils are the front line of defense against S. aureus infections. By modulating neutrophil behavior and inducing premature cell death, S. aureus has an advantage during an infectious state. In this thesis, I provide a method for studying this host and pathogen dynamic, and moreover, I investigate the mechanism by which S. aureus inhibits the neutrophil inflammatory response by repressing NF-KappaB. Here I show that S. aureus secretes a protein 30-50kDa in size, which both decreases total amount of NF-KappaB and activated NF-KappaB in neutrophils. This potent mystery protein is able to repress IL-8 production and does this all in a lysis independent manner. Additionally, the mystery protein is able to inhibit NF-KappaB activity in another cell type, the monocyte. It was previously believed that the S. aureus protein SSL3 was responsible for deactivating NF-KappaB, but herein, I show this is not the case. These findings reopen the need to examine the mechanism by which S. aureus modulates neutrophil inflammatory responses. Inhibition of the inflammatory response is likely linked the premature cell death seen during S. aureus infections. By utilizing these clues, the field is closer to understanding the intricacies of this host and pathogen dynamic, opening avenues to developing novel infection treatment methods.Item Nurse-driven catheter-associated urinary tract infection (CAUTI) prevention project(Montana State University - Bozeman, College of Nursing, 2021) Gaskin, Katelyn Dawn; Chairperson, Graduate Committee: Susan LuparellAn indwelling urinary catheter (IUC) is a tube that is placed into the bladder through the urinary tract, left in place and connected to a closed system drainage bag. Catheter placement in the urinary tract increases the risk of bacteria ascending the catheter and causing an infection, known as a catheter-associated urinary tract infection (CAUTI) (Center of Disease Control [CDC], 2019; Fekete, 2020). Healthcare-associated infections (HAI) are the most common complication of healthcare treatment and are a major cause of mortality and morbidity. CAUTIs are the most common and preventable HAI, making up over 30% of the HAI in the United States (Agency of Healthcare Quality and Research [AHRQ], 2015; CDC, 2019; Centers for Medicare & Medicaid Services [CMS], 2008). CAUTIs cause increase pain and discomfort, and increase patients' hospital length of stay (AHRQ, 2017; CDC, 2019). IUCs are frequently placed without appropriate indication and remain in longer than medically necessary (CDC, 2019). Problem statement: Critical care patients are most vulnerable to acquiring a CAUTI from an IUC due to their weakened immune systems and underlying co-morbidities (CDC, 2019). The intensive care unit (ICU) setting has the highest reported rates of CAUTIs (CDC, 2019). Purpose statement: The purpose of this project was to reduce CAUTIs by implementing a nurse-driven algorithm to avoid IUC insertion, decrease IUC duration, and strengthen existing CAUTI prevention measures. Methods: Kotter's change management theory combined with the Plan-Do-Study-Act (PDSA) model served as the project's framework. The project was designed to foster a collaborative approach to reduce CAUTI incidences by empowering nurses to work at the highest level of their scope of practice, standardizing care, and strengthen existing CAUTI prevention. Results: CAUTIs were measured pre/post-implementation, and an absolute reduction from 2 to 0 was observed. The standard infection ratio (SIR) decreased to 0 post-implementation and the standard utilization ratio (SUR) decreased by 2.17%. ICU nurses were very likely (92%) to implement the algorithm into practice, and 81% indicated they implemented the algorithm on 75% of their patients. Conclusion: Although many studies have looked at reducing CAUTIs and have shown the benefit of avoiding insertion and using alternatives, there are few that have examined the standardization and combination of alternative measures, appropriate indications, and acute urinary retention measures into one nurse-driven algorithm. This quality improvement project implemented evidence-based practice in a nurse-driven algorithm and observed a decrease in CAUTI incidence.Item Assessment of variation in the detection and prevalence of blood parasites among sympatrically breeding geese in western Alaska(Montana State University - Bozeman, College of Letters & Science, 2021) Buchheit, Raymond Matthew; Chairperson, Graduate Committee: Jay J. Rotella; Joel A. Schmutz, Brian Uher-Koch and Andrew M. Ramey were co-authors of the article, 'Assessment of variation in the detection and prevalence of blood parasites among sympatrically breeding geese in western Alaska' submitted to the journal 'Journal of wildlife diseases' which is contained within this thesis.Haemosporidian parasites may impact avian health and are subject to shifts in distribution and abundance with changing ecological conditions. Therefore, understanding variation in parasite prevalence is important for evaluating biologically meaningful changes in infection patterns and associated population level impacts. Previous research in western Alaska indicated a possible increase in Leucocytozoon infection between emperor geese (Anser canagicus) sampled in 1996 (<1%, n=134), and during 2011-2012 (19.9%, 95% CI: 3.0-36.8%, n=77); however, different detection methods were used for these estimates. Prior research in this same region identified a lack of Leucocytozoon parasites (0%, n=117) in sympatrically breeding cackling geese (Branta hutchinsii minima) in 2011. In this study, we molecularly screened blood samples collected from sympatrically breeding emperor and cackling geese in western Alaska during additional breeding seasons to better assess temporal and species-specific variation in the prevalence of blood parasites. We found similar prevalence estimates for Leucocytozoon parasites in emperor goose blood samples collected in 1998 and 2014, suggesting consistent infection of emperor geese with blood parasites at these time points. Using samples from sympatric geese sampled during 2014, we found evidence for higher incidence of Leucocytozoon parasites among emperor geese (20.3%, 95% CI: 11.8-32.7%) as compared to cackling geese (3.6%, 95% CI: 1.1-11.0%) reinforcing the previous finding of species-specific differences in infection. Furthermore, we detected Leucocytozoon, Haemoproteus, and Plasmodium blood parasites in unflighted goslings of both species, supporting the possible transmission of these parasites at western Alaska breeding grounds. Our results help to clarify that prevalence of Leucocytozoon parasites have likely remained consistent among emperor geese breeding in western Alaska since the late 1990s and that this species may disproportionally harbor Leucocytozoon parasites as compared to sympatrically breeding cackling geese.