Year

2020

Degree Name

Doctor of Philosophy

Department

School of Chemistry and Molecular Biosciences

Abstract

Group A Streptococcus (GAS) is a major bacterial causative agent of a wide variety of human diseases that range from mild superficial infections such as impetigo, and pharyngitis, to more serious invasive infections, and numerous autoimmune sequelae. Globally, GAS is estimated to cause 700 million infections, and accounts for half a million deaths per year, and consequently poses a considerable burden financially and on healthcare systems. Of the GAS disease etiologies, GAS pharyngitis is the most common disease manifestation, with an annual estimate of 616 million cases globally. Penicillin is the standard antibiotic treatment of GAS infections. Whilst GAS remains sensitive to penicillin, an alarming antibiotic treatment failure rate of 20-40% has been reported. It has been proposed that GAS may exist as biofilms in the pharynx contributing to persistent and recurrent pharyngitis that is non-responsive to antibiotic treatment. In vitro, plate-based models have shown that several GAS M-types form biofilms, and multiple GAS virulence factors have been linked to biofilm formation. Although the contributions of these plate- based studies have been valuable, most have failed to mimic the host environment, with many studies utilising abiotic surfaces. Moreover, GAS is a human specific pathogen, and colonisation and subsequent biofilm formation is likely facilitated by distinct interactions with host tissue surfaces. Thus, the overarching aim of chapter 2 was to optimise a GAS biofilm-host pharyngeal cell model to support and grow GAS biofilms of a variety of GAS M-types. Improvements and adjustments to the crystal violet biofilm biomass assay were also tailored to reproducibly detect delicate GAS biofilms. 72 h was deemed as an optimal growth period for yielding detectable biofilm biomass. The GAS biofilms formed were robust and durable, and can be reproducibly assessed via staining/washing intensive assays such as crystal violet with the aid of methanol fixation prior to staining. SEM imaging of GAS biofilms formed by this model resembled those previously found on excised tonsils of patients suffering chronic pharyngo-tonsillitis. Taken together, an efficacious GAS biofilm host-cell model has been developed that can support long-term GAS biofilm formation, with biofilms formed closely resembling those seen in vivo.

Share

COinS
 

Unless otherwise indicated, the views expressed in this thesis are those of the author and do not necessarily represent the views of the University of Wollongong.