Natural and mimetic vesicles to elicit a potent innate immune response against MRSA infection

• Main Author: Ong, Samuel Vernon Jing Heng
• Other Authors: Czarny Bertrand Marcel Stanislas
• Format: Final Year Project
• Language: English
• Published: Nanyang Technological University 2024
• Subjects: Medicine, Health and Life Sciences; Staphylococcus aureus; Bacterial membrane vesicles; Mimetic vesicles; Macrophage activation; qPCR
• Online Access: https://hdl.handle.net/10356/176160
• Institution: Nanyang Technological University

Staphylococcus aureus (S. aureus or SA) is a widespread bacterial pathogen that can cause serious infections that can lead to life-threatening conditions. Methicillin-resistant SA (MRSA) is a strain of SA that has generated resistance to front-line antibiotics and is significantly harder to treat due to MRSA clones having the ability to acquire resistance to new drugs and anti-microbial agents quickly. MRSA clones are also specific to different world regions and have varying virulence mechanisms, which presents a challenge in employing effective treatment methods for patients suffering from MRSA infections. The lack of an effective and licensed vaccine is the motivation for studies into novel vaccines, which can hopefully present a way to effectively prevent MRSA infections and reduce the burden on MRSA treatment methods. This project aims to explore the potential for extracellular vesicles (EVs) produced from SA and MRSA bacterial cultures to elicit a potent innate immune response from immune cells against MRSA infections, thereby supporting the novel idea of utilising MRSA EVs to produce vaccines for MRSA infections. In this project, natural EVs (NEVs) were extracted from SA and MRSA bacterial cultures. Mimetic EVs (MEVs) were isolated from SA and MRSA protoplasts through a spin-cup method, and purification of NEVs and MEVs was done using ultra-centrifuge and size-exclusion chromatography (SEC). Isolated EVs were characterised through Bicinchoninic acid (BCA) protein assay and nanoparticle tracking analysis (NTA) and used to treat RAW264.7 macrophage cells. The cytokine gene expression of activated macrophages was analysed using real-time quantitative polymerase chain reaction (RT-qPCR). Results showed that EVs, especially MRSA MEVs, were able to elicit appropriate cytokine expressions by macrophages, which implies that MRSA MEVs can trigger an immune response by host cells to develop immunity against MRSA infections and supports the potential for MRSA MEVs to be further studied as a vaccine.