New CRISPR-Based Tool Enables Genome-Wide Mutagenesis of Bacteriophages

Researchers at the University of Otago in New Zealand have developed a CRISPR-based method for systematically disrupting genes across bacteriophage genomes. This approach, detailed in the study "Defining the essential genome of diverse phages with phage Tn-seq," published in Nature Microbiology, aims to expedite the process of understanding gene functions and engineering phages for therapeutic and biotechnology applications.

Bacteriophages, or phages, are viruses that infect bacteria and have shown promise in addressing antimicrobial resistance and reducing reliance on agrochemicals. However, many phage genes encode unknown functions, complicating predictions of viral behavior and reliable modification. The research team combined transposon insertion sequencing with CRISPR–anti-CRISPR selection to overcome these challenges.

This technique involves inserting a transposon into a phage genome to disrupt a gene, followed by a selection system to recover phages carrying those mutations, thereby identifying essential versus non-essential genes. Building on this mutagenesis workflow, the researchers expanded the method to introduce new genetic cargo, such as fluorescent markers or genes that help phages overcome bacterial defense systems.

This dual capability allows for both the mapping of phage genomes and the rapid engineering of phages with new functions. Senior author Peter Fineran highlights the significance of this work in bridging the gap in our understanding of phages, similar to the initial stages of antibiotic research in the 1950s. The platform's broad applicability and cost-effectiveness make it a promising tool for both fundamental studies and the development of novel phage-based therapies.

Nonetheless, engineered phages would still require rigorous testing to ensure their safety and efficacy in practical applications.