Phagemid-based capsid system for CRISPR-Cas13a antimicrobials targeting methicillin-resistant Staphylococcus aureus

Area of Science:

• Biotechnology
• Molecular Biology
• Antimicrobial Resistance

Background:

• Escalating antibiotic resistance in multidrug-resistant pathogens poses a significant global health threat.
• Targeted therapies are needed to combat resistant bacterial infections like Staphylococcus aureus.

Purpose of the Study:

• To develop an innovative phagemid-based capsid system for generating CRISPR-Cas13a-loaded antibacterial capsids (AB-capsids).
• To optimize the system for efficient production and targeted therapy against multidrug-resistant Staphylococcus aureus.

Main Methods:

• Optimization of a phagemid system to maximize AB-capsid yield and purity.
• Utilizing prophage-encoded packaging site genes (terL-terS-rinA-rinB) for high packaging efficiency.
• Employing lysogenized hosts with terL-terS deletion to minimize wild-type phage contamination.

Main Results:

• A positive correlation was observed between phagemid copy number and AB-capsid yield, with an 8.65-fold increase in copy number leading to a 2.54-fold rise in AB-capsid generation.
• Phagemids with terL-terS-rinA-rinB genes showed high packaging efficiency.
• AB-capsid generation in hosts with terL-terS deletion reduced wild-type phage contamination with minimal impact on yield.

Conclusions:

• The developed phagemid-based capsid system is a promising approach for producing sequence-specific bactericidal agents.
• This system offers an efficient and targeted strategy to combat antibiotic-resistant pathogens.