New tool to mutate phages boosts the fight against superbugs

A new tool to mutate bacteriophages – viruses that infect bacteria – provides a “big leap” forward in both our understanding of how they function, and our ability to harness their full potential.

Developed by researchers in the University of Otago – Ōtākou Whakaihu Waka, New Zealand, Department of Microbiology and Immunology, the method features on the latest cover of prestigious international journal Nature Microbiology.

Senior author Professor Peter Fineran says bacteriophages, or simply phages, have “huge potential” to combat the antimicrobial resistance crisis, and for increasing sustainable agriculture as an alternative to agrochemicals, because they can destroy bacteria.

“But our knowledge of phages is probably like the understanding of antibiotics back in the 1950s. Many phage genes are currently in the area of microbial dark matter – encoding functions we just don’t understand – which is limiting our ability to use phages in healthcare and biotechnology.

“This new development is a big leap in how we can rapidly understand phage biology and then use that knowledge to make phages work better to help achieve our goals.” 

Co-lead author Dr Manuela Fuchs says the new method allows for genome-wide mutagenesis of bacteriophages using CRISPR-Cas technology.

The researchers utilised transposon insertion sequencing – using a mobile piece of DNA which can jump in and disrupt a gene – combined with CRISPR-anti-CRISPR-based selection, to select phages that have been mutated. This allows them to identify genes that are essential and genes that are non-essential for phage survival.

Once we had that established, we thought we could expand the technique and also use it to add genes directly into the phage genomes. We essentially found it is possible to load additional genes on that transposon, not just our anti-CRISPR protein. While we added a fluorescent marker, it could be used to, for example, add additional anti-defence genes to phages, to improve their therapeutic potential.” 

Dr. Manuela Fuchs, Co-lead author, University of Otago 

Senior author Dr Leah Smith says the method enables genes to be automatically inserted somewhere while maintaining phage function.

“This is a systematic, broadly applicable, and cost-effective way to not only investigate gene function, but also rapidly engineer phage genomes.

“This opens up new opportunities for both fundamental research and future therapeutic development.”

One particular example of this would be biofilms, which can cause hard-to-treat infections on prosthesis implants and medical devices.

“With this new technique, we could quickly load phages to have more things that stop some bacterial defences so that they can be harnessed to kill pathogens more easily,” Dr Smith says.