Development of Pneumolysin as a vaccine candidate

Mughal, Muhammad Kashif (2014) Development of Pneumolysin as a vaccine candidate. PhD thesis, University of Glasgow.

Abstract

More than 90 serotypes of Streptococcus pneumoniae are responsible for causing a number of invasive and non-invasive diseases in humans. To combat this pathogen there are two types of vaccines available namely pneumococcal polysaccharide vaccine (PPV) and pneumococcal conjugate vaccine (PCV). Although these vaccines have reduced the incidence and prevalence of pneumococcal diseases yet these vaccines are unable to curb the disease and have some potential drawbacks. The problems associated with PPV is inability to mount appropriate immune response in young children (<2 years) of age and with PCV, disadvantages are serotype replacement and affordability. These disadvantages have shifted the focus of attention on developing vaccines that can provide serotype independent protection and is cost effective and can be used in developing and under-developing countries as well.
Pneumolysin (PLY) is one of the members of cholesterol dependent cytolysin (CDC) family and is one of the key virulence factors of the bacterium. PLY has a wide variety of functions but the two most important functions are the ability to lyse the host cell by forming pores and activating the complement pathway. Studies have shown recombinant PLY to be a good vaccine candidate; however PLY is toxic and cannot be used as a vaccine. A number of toxoid versions have been made targeting residues responsible for toxin’s lytic activity and tested in-vivo models. ∆6 PLY is a toxoid made by deleting two amino acid residues and seems to be a potential candidate to be included in next generation pneumococcal vaccines. ∆6 PLY is immunogenic and act as an adjuvant as well, however ∆6 PLY causes the aggregation of red blood cells and therefore cannot be used in the vaccine.
The present study is carried out to identify the residue/s responsible for causing this agglutination behaviour. A number of molecular biology techniques were used in this study for making mutants in different backgrounds and were tested for their lytic and agglutination activity. The study identified the single residue in domain 4 causing aggregation of the red blood cells. The agglutination negative mutant was then tested at different concentrations through a series of experiments including hemagglutination assays, fluorescence microscopy and fluorescence activated cell sorter (FACS).
The study also highlighted some important regions of the toxins responsible for maintaining structure of the toxin. This new toxoid seems to have the potential to be used in future pneumococcal vaccines alone, in combination with other pneumococcal proteins and/or polysaccharides.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: Q Science > QR Microbiology; R Medicine > RZ Other systems of medicine
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Infection & Immunity > Bacteriology
• Supervisor's Name: Mitchell, Professor T.J.
• Date of Award: 2014
• Depositing User: Muhammad Kashif Mughal
• Unique ID: glathesis:2014-5151
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 05 May 2014 10:06
• Last Modified: 12 Jul 2017 13:14
• URI: https://theses.gla.ac.uk/id/eprint/5151