Morck, Douglas WalterLockhart, Joey Scott2022-05-102022-05-102022-05-06Lockhart, J. S. (2022). Biofilm-derived Small Molecules Released from Mixed-species Cultures of Fusobacterium necrophorum and Porphyromonas levii Inhibit Bovine Neutrophil Functional Responses (Doctoral thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca.http://hdl.handle.net/1880/114649Bacterial biofilms are surface adhered communities of microorganisms encased in a protective polysaccharide layer. These resilient structures present significant challenges for effective host immune cell clearance or clinical interventions, frequently resulting in recurrent infections in cattle, humans, and other animals. Fusobacterium necrophorum and Porphyromonas levii are two species of anaerobic, Gram-negative, opportunistic pathogens that have been described as etiological agents in a variety of necrotic, recalcitrant infections. With antimicrobial resistance on the rise, it is important to elucidate the reasons why biofilm-associated infections are so difficult to eradicate. Neutrophils are critical first responder cells that are recruited to a site of bacterial colonization to initiate the inflammatory response. In this dissertation research, an in vitro system for the growth of mixed-species anaerobic biofilms was employed to investigate bovine neutrophil responses to biofilm bacteria compared to planktonic cultures of the same two species. Neutrophils exposed to biofilm soluble factors generated reduced oxidative and chemotactic responses compared to neutrophils exposed to planktonic bacteria-generated products. Ultra-filtration revealed that a <3kDa molecule is differentially expressed in the bacterial supernatants and allows biofilm-bacteria to inhibit neutrophil responses. Intensive heat, protease, lipase, or nuclease treatment of the <3kDa fractions prior to exposure to bovine neutrophils did not alter the differential oxidative responses. This led to an investigation of small, heat stable compounds and our results suggest that the accumulation of protoporphyrin IX in the biofilm mode of growth attenuates neutrophil chemotaxis and reactive oxygen species production. This work provides evidence for a mechanism by which these biofilm bacteria evade host immune cells and understanding such mechanisms is important to reduce the persistent inflammation and negative clinical outcomes associated with chronic biofilm-mediated disease.engUniversity of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.Education--Sciencesdoctoral thesishttp://dx.doi.org/10.11575/PRISM/39764