The investigation of novel virulence genes in Porphyromonas gingivalis

This study focuses on Porphyromonas gingivalis, a Gram negative anaerobic bacterium and a “Keystone” pathogen involved with the progression of periodontal diseases. P. gingivalis produces multiple virulence factors that have been linked to other chronic diseases such as cardiovascular disease and Alzheimer’s disease. Through the use of fimbriae, gingipain proteases, lipopolysaccharide, and outer membrane proteins, this bacteria has the ability to interact with its host. These host-pathogen interactions allow P. gingivalis to bind and invade host cells, influence an inflammatory response whilst simultaneously suppressing local immune defences. This provides a better niche for the survival of the pathogenic subgingival microbiota that progress the chronic inflammatory conditions of periodontal disease. In this thesis, the virulence of P. gingivalis is investigated through the study of three genes indicated to be involved in the invasion of host cells, ompH1, rfbB and PGN2012, whilst simultaneously attempting to identify human receptors involved with P. gingivalis adhesion.
Through knockout mutagenesis, these genes have been disrupted and the characterisation of these mutants through investigating their virulence has been assessed. The outer membrane protein chaperone OmpH1 mutant resulted in reduced adhesion and invasion of H357 (oral epithelial) cells, and reduced protease activity, providing more evidence that the ompH chaperone proteins have clients involved in host-pathogen interactions. The rhamnose synthesis gene mutant rfbB produces a truncated lipopolysaccharide and has demonstrated a decreased virulence in multiple assays including biofilm formation, adhesion and invasion of H357 cells, and protease activity. Finally, it has been demonstrated that the efflux channel protein gene PGN2012 is involved with heavy metal efflux, specifically Zn2+, Co2+ and Cd2+, whilst having no significant role in adhesion and invasion of H357 cells. This indicates that PGN2012 may have a role in intracellular survival rather than the mechanics of invading host cells.
Overall, we demonstrate the important role of surface components in the host-pathogen interactions of P. gingivalis. This work also reveals that the PGN2012 operon is the CzcCBA heavy metal efflux system in P. gingivalis.