Theses and Dissertations at Montana State University (MSU)
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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 Characterization of host-pathogen interactions during early Staphylococcus aureus biofilm formation on surfaces(Montana State University - Bozeman, College of Agriculture, 2022) Pettygrove, Brian Alexander; Chairperson, Graduate Committee: Philip S. Stewart; This is a manuscript style paper that includes co-authored chapters.Implanted biomaterials such as orthopedic screws, prosthetic joints, pacemakers, and catheters are essential components of modern medicine. Unfortunately, implanted foreign bodies are susceptible to biofilm infection, leading to a persistent and difficult to treat disease state. Biofilm infections readily tolerate clearance from the immune system, however much of our understanding of the mechanisms governing persistence are formulated around the biofilm state during advanced infection. By comparison we have a poor understanding of the early stages of infection. Specifically, how contaminating organisms initially evade host immune defenses and establish a robust infection remains ill-defined. In this work, we interrogated interactions between Staphylococcus aureus (S. aureus), a frequent culprit in biomaterial infections, and early contributors to host immunity. Using in vitro time-lapse microscopy, we observed that human neutrophils readily phagocytose and kill single cells or small clusters of S. aureus cells that are attached to a surface. S. aureus cells that go undiscovered during the initial stages of neutrophil surveillance form biofilm aggregates that rapidly gain tolerance to neutrophil killing. In vivo models of implant infection demonstrated that surface adherent bacteria can evade discovery due to delayed or heterogeneous neutrophil recruitment to the surface. Biofilm aggregate formation was impaired in a strain deficient in the two-component gene regulatory system SaeR/S and the resulting cells were highly susceptible to neutrophil killing. Inhibition of aggregation was dependent on serum complement proteins C3 and factor B, suggesting that SaeR/S regulated factors actively inhibit host complement to facilitate aggregation. Taken together, these data suggest that the formation of immune-tolerant biofilm aggregates may contribute to chronic device related infections by protecting bacteria from phagocyte killing. These studies provide vital insight into the host pathogen interactions on contaminated biomaterial surfaces and highlight early events that may determine infection outcome.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.