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Matthew J Pestrak Thesis.pdf (4.29 MB)

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Investigation of the Pseudomonas aeruginosa biofilm exopolysaccharide Psl and its role during infection

Pestrak, Matthew James, Pestrak
http://rave.ohiolink.edu/etdc/view?acc_num=osu1543240479329587

Abstract Details

2018, Doctor of Philosophy, Ohio State University, Biomedical Sciences.
P. aeruginosa is one of the most common causes of nosocomial infection and is frequently the cause of lethal lung and wound infections. These infections are often difficult to treat due to the formation of aggregated communities, known as biofilms. The process of biofilm formation is complex and involves the production and secretion of a protective extracellular matrix. Once encased in the biofilm, P. aerguinosa becomes more tolerant to killing by antimicrobials, host immune factors, and other environmental stressors. Thus, these infections are often difficult to treat and frequently lead to chronic infection. A major component of the biofilm matrix is the exopolysaccharide (ePS) Psl. This exopolysaccharide provides structural support for the biofilm and aids in cell-cell and cell-surface attachment. Additionally, Psl protects bacteria and promotes drug tolerance by sequestering antimicrobials and immune components before they reach the active bacterial population. Over the course of infection, P. aeruginosa adapts to the host environment leading to mutations which promote survival. Strains with mutations causing overproduction of the ePS, Psl and Pel, are frequently isolated from chronic infection. These hyperbiofilm forming strains have been named rugose small colony variants (RSCVs), and their presence has been correlated to poor prognosis. A positive selective pressure for RSCVs exemplifies the importance of Psl during infection, but it was unclear specifically what ePS overproduction does during infection. Here, we describe how Psl overproduction negatively effects the host. Considering these effects, we then sought to 1) target Psl directly with therapeutics and 2) identify novel Psl regulators to provide new targets for disrupting synthesis. We report that the overproduction of Psl by RSCVs leads to auto-aggregation and inflammation in the lung. The clustering of bacterial cells promotes host recognition resulting in ROS and pro-inflammatory cytokine production. The excessive inflammation had devastating effects for the host leading to significant tissue damage and delayed wound healing. Despite a robust inflammatory response, RSCVs exhibited enhanced evasion of phagocytosis resulting in greater bacterial burden during infection. These data provide insight on the detrimental effects of Psl overproduction and bacterial aggregation during infection. Given the antagonistic role of Psl, we sought a novel way to treat P. aeruginosa infection by degrading Psl with the glycosyl hydrolase PslG. We demonstrate PslG potentiates the killing activity of a number of antibiotics, and it improves tobramycin bacterial clearance during wound infection. Finally, the process of Psl production is a complex and energy intensive process that must be tightly regulated. All of the factors involved in the regulation of Psl remains unclear. Here we utilized a variety of screening methods to search for new potential Psl regulators, providing the groundwork for identifying new Psl disrupting therapeutic targets. Collectively, our findings here contribute to furthering our understanding of Psl regulation and its role during infection. This work gives us new insights on the role of aggregation as a virulence mechanism. Additionally, we have identified a new strategy for managing P. aeruginosa infection, and these findings demonstrate that future work should be done to develop therapeutics to disrupt the biofilm matrix and exopolysaccharides during infection.
Daniel Wozniak (Advisor)
Stoodley Paul (Committee Chair)
Gunn John (Committee Member)
Mason Kevin (Committee Member)
195 p.
Biomedical Research; Microbiology
Pseudomonas aeruginosa, biofilm, exopolysaccharide, Psl, rugose small colony variant, wound infection, lung infection, host-pathogen interactions

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Citations

  • Pestrak, Pestrak, M. J. (2018). Investigation of the Pseudomonas aeruginosa biofilm exopolysaccharide Psl and its role during infection [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1543240479329587

    APA Style (7th edition)

  • Pestrak, Pestrak, Matthew. Investigation of the Pseudomonas aeruginosa biofilm exopolysaccharide Psl and its role during infection. 2018. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1543240479329587.

    MLA Style (8th edition)

  • Pestrak, Pestrak, Matthew. "Investigation of the Pseudomonas aeruginosa biofilm exopolysaccharide Psl and its role during infection." Doctoral dissertation, Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1543240479329587

    Chicago Manual of Style (17th edition)

osu1543240479329587
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This open access ETD is published by The Ohio State University and OhioLINK.