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Published on: April 11, 2019
Phagemid-based capsid system for CRISPR-Cas13a antimicrobials targeting methicillin-resistant Staphylococcus aureus
Feng-Yu Li1, Xin-Ee Tan1, Yuzuki Shimamori1
1Division of Bacteriology, Department of Infection and Immunity, School of Medicine, Jichi Medical University, Shimotsuke city, Tochigi, 329-0498, Japan.
Abstract:
In response to the escalating antibiotic resistance in multidrug-resistant pathogens, we propose an innovative phagemid-based capsid system to generate CRISPR-Cas13a-loaded antibacterial capsids (AB-capsids) for targeted therapy against multidrug-resistant Staphylococcus aureus. Our optimized phagemid system maximizes AB-capsid yield and purity, showing a positive correlation with phagemid copy number. Notably, an 8.65-fold increase in copy number results in a 2.54-fold rise in AB-capsid generation. Phagemids carrying terL-terS-rinA-rinB (prophage-encoded packaging site genes) consistently exhibit high packaging efficiency, and the generation of AB-capsids using lysogenized hosts with terL-terS deletion resulted in comparatively lower level of wild-type phage contamination, with minimal compromise on AB-capsid yield. These generated AB-capsids selectively eliminate S. aureus strains carrying the target gene while sparing non-target strains. In conclusion, our phagemid-based capsid system stands as a promising avenue for developing sequence-specific bactericidal agents, offering a streamlined approach to combat antibiotic-resistant pathogens within the constraints of efficient production and targeted efficacy.
Insights
This study introduces a novel system using bacteriophage capsids loaded with CRISPR-Cas13a to combat antibiotic-resistant bacteria, specifically targeting Staphylococcus aureus. The optimized system enhances production efficiency and ensures targeted bacterial elimination.
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.
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