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Title: Investigation of the sialidases of periodontal pathogens
Author: Frey, Andrew
ISNI:       0000 0004 6062 5723
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2017
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Periodontitis results in destruction of tooth supporting structures, eventually leading to tooth loss, and it affects ~10% of the world's population. Key to its onset and progression is a complex relationship between the periodontal bacteria and the host inflammatory response. The bacteria most associated with severe periodontitis are the so-called periodontal pathogens of the red complex- Tannerella forsythia, Treponema denticola, and Porphyromonas gingivalis. These organisms all express sialidases, which cleave sialic acid from host glycoproteins, and this is believed to contribute to disease. Considering this, the aims of this project were to further characterise the sialidases of T. forsythia and P. gingivalis (NanH and SiaPG), to test their inhibition with commercially available chemotherapeutics, and the effect this has on in vitro models of virulence. NanH and SiaPG were successfully purified by affinity chromatography. The enzymes were shown to possess a high degree of activity over various conditions and on different sialic acid ligands. Importantly, both sialidases could be inhibited using the commercially available chemotherapeutic zanamivir (supplied by GlaxoSmithKline, UK) which laid the groundwork for studies testing inhibition of virulence. The accessory enzyme NanS was shown to bolster sialic acid release from bovine salivary mucin by both NanH and SiaPG. This was important for nutrient acquisition, since NanS also enhanced the growth of P. gingivalis in defined media where serum and saliva were the only nutrient sources. These findings also hinted at the possibility of interspecies cooperation in sialic acid release from host sources. Zanamivir also inhibited biofilm formation of P. gingivalis on oral-relevant glycoprotein sources. Zanamivir was also shown to be capable of inhibiting attachment and invasion of oral epithelial cells by P. gingivalis, T. forsythia, and the sialidase negative pathogen Fusobacterium nucleatum, even when multiple species were present during infection. Finally NanH was shown to part-mediate pro-inflammatory signalling in oral epithelial cells in response to P. gingivalis LPS, and zanamivir prevented pro-inflammatory cytokine release in cells infected with T. forsythia. This highlighted multiple mechanisms by which sialidase inhibition can prevent host-pathogen interactions. This study broadens our understanding of the multifarious roles of bacterial sialidases in virulence, and indicates that their inhibition with chemotherapeutics could be a promising strategy for periodontitis therapy.
Supervisor: Stafford, Graham ; Murdoch, Craig ; Charles, Douglas Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available