Production And Characterization Of Biological Nanoparticles For The Development Of A Novel Vaccine Against Methicillin-resistant Staphylococcus Aureus
Methicillin-resistant Staphylococcus aureus (MRSA) is a Gram-positive bacterium which poses a substantial threat to public health due to its high virulence and multi-drug resistance. Membrane vesicles (MVs) are naturally-derived nanoparticles which have been shown to provide protection against bacterial infections. In this study, MVs were investigated as a potential vaccination strategy against MRSA infection. MVs were extracted from a USA300 strain of MRSA and visualized with transmission electron microscopy. Antisolvent crystallization was found to be a more efficient extraction method than ultrafiltration, and MVs were successfully purified from solution using ammonium sulfate precipitation. Protein content, particle size, and dispersity in solution were studied using gel electrophoresis and dynamic light scattering. MV samples were found to be within 20-200 nm in diameter and to aggregate substantially in solution. MV preparations were tested for toxicity in vitro with RAW 264.7 murine macrophages and in vivo with a Galleria mellonella model. MRSA-derived MVs demonstrated lower toxicity than E. coli MVs, even at higher concentrations. G. mellonella treated with MRSA MVs had a 70% survival rate through 5 days, compared to a 10% survival rate in the E. coli MV group. Protection against infection was investigated in BALB/c mice, which showed IgG responses specific for MRSA after two intranasal immunizations. Mice were challenged intranasally with 1x108 CFU of MRSA and monitored for morbidity and mortality. Mice immunized with MVs displayed significantly lower bacterial burdens in their lungs compared to naÃ¯ve mice. The results of these experiments indicate that MRSA MVs may represent a novel vaccine strategy for preventing MRSA infection.