Phage in the Age of Synthetic Biology

By Karen Weynberg, CSIRO Synthetic Biology Fellow, School of Chemistry and Molecular Biosciences, Faculty of Science, University of Queensland

Synthetic biology is an emerging field of research set to revolutionise the future of biological science. To simply summarise synthetic biology can be challenging due to the myriad of tools, techniques and applications it encompasses. In essence, synthetic biology involves the use of engineering principles in a biological context. Using DNA-encoded componentry, synthetic biology enables the design and construction of biological parts, devices, cellular circuits and networks, and even whole organisms. This exciting new approach holds great promise for many areas of research, including biomedical efforts to treat cancer and other diseases, vaccine development, cell therapies, regenerative medicine and microbiome engineering.

The growing problem of global antibiotic resistance is another area that synthetic biology can play a crucial role in. The abuse and misuse of antibiotics in human health and animal agriculture has resulted in an alarming increase in multi-drug resistant (MDR) bacterial strains. Future projections predict that antibiotic resistance could result in as many as 10 million deaths a year by 2050, surpassing mortality rates due to cancer¹. Alternatives to antibiotics are now being sought, including phage therapy that is based on the natural mechanisms by which bacteriophages (literally ‘bacteria eaters’) infect and kill bacteria.  

Phage therapy can be encapsulated by the ancient proverb that states ‘The enemy of my enemy is my friend’.

Phages were discovered just over 100 years ago by two independent European researchers, Frederick Twort and Felix D’Herelle. Since then, phage therapy has been used most notably in Eastern European countries, such as Russia and Georgia, to treat bacterial infections in human patients. The discovery of antibiotics led Western medicine to turn away from phage therapy, but this cheap, natural and targeted treatment is now increasingly appealing. Phage therapy is poised to be the ‘Next Big Thing’ in fighting deadly superbug infections, as the recent increase in both fundamental research and interest from biotech companies attests to. Companies such as US-based Ampliphi Biosciences, China-based Phage Luxe and France-based Pherecydes Pharma are directing efforts into developing phage therapy as a viable alternative to antibiotic treatments.

Typically, a cocktail comprised of a number of different phages is administered to combat a bacterial infection. Phage are usually highly specific for a bacterial host so the unwanted side effects of broad-spectrum antibiotics, such as targeting beneficial bacteria not just pathogens, are avoided in phage therapy. Although, hurdles are still to be overcome, including the approval of rigorous advanced clinical trials, there have been incidences of ‘compassionate cases’ successfully treated with phage therapy. One such case is described by Dr Steffanie Strathdee in her TEDx talk on how phage saved her husband’s life from an otherwise fatal MDR infection – a case that SIG member Dr. Jeremy J. Barr directly contributed to:

Link to Dr Steffanie Strathdee TEDx talk:

https://www.youtube.com/watch?v=AbAZU8FqzX4

Synthetic biology can be used to engineer and synthesise phages to increase their natural capabilities and manipulate their host range. Issues arising from the use of traditional phage therapy, such as bacterial resistance and limited host range can be overcome by employing synthetic biology techniques. Wide-ranging approaches to engineering phage include in vivo recombineering, CRISPR-Cas mediated genome engineering, whole-genome synthesis and cell-free transcription-translation systems (for a comprehensive review see Pires et al. 2016²). Yeast-based assembly of phage genomes also enables the refactoring of phage genomes^3,4. Phage genomes can be redesigned in silico based on natural phage genome sequence templates. Phage genome PCR products are then captured in a yeast artificial chromosome (YAC) and propagated within the yeast cell, which acts an intermediate host, thus avoiding toxicity issues that would arise in a bacterial host. Once assembled the engineered phage genomes are extracted and electroporated into a bacterial host, in order to reboot the phage and produce viral progeny.

Pathogenic bacteria can persist within a biofilm that is comprised of a multi-species consortium within a dense extracellular matrix. Biofilms can form within medical devices, such as catheters leading to health problems including, for example, urinary tract infections. The formation of biofilms helps bacteria to evade the host immune response and minimise their exposure to antibiotics. Phage can be engineered to produce enzymes e.g. dispersins⁵ to degrade biofilms, thereby making target bacteria vulnerable to phage lysis, antibiotics and immune responses such as macrophages. Engineering of phage to produce other degrading enzymes, such as capsule depolymerases and lysins, can also increase the efficiency of phage infection.

A recent review comprehensively describes the wide scope of phage-based applications that are now feasible using the sophisticated techniques and tools afforded by synthetic biology⁶. This field of biological research is set to greatly accelerate the development of novel phage-based solutions to a wide range of issues. The future is bright, the future is phage!

References

1          Editors, P. M. Antimicrobial Resistance: Is the World Unprepared? PLoS medicine 13, e1002130-e1002130, doi:10.1371/journal.pmed.1002130 (2016).

2          Pires, D. P., Cleto, S., Sillankorva, S., Azeredo, J. & Lu, T. K. Genetically Engineered Phages: a Review of Advances over the Last Decade. Microbiology and Molecular Biology Reviews 80, 523 (2016).

3          Ando, H., Lemire, S., Pires, D. P. & Lu, T. K. Engineering Modular Viral Scaffolds for Targeted Bacterial Population Editing. Cell Systems 1, 187-196, doi:10.1016/j.cels.2015.08.013 (2015).

4          Jaschke, P. R., Lieberman, E. K., Rodriguez, J., Sierra, A. & Endy, D. A fully decompressed synthetic bacteriophage øX174 genome assembled and archived in yeast. Virology 434, 278-284, doi:10.1016/j.virol.2012.09.020 (2012).

5          Lu, T. K. & Collins, J. J. Dispersing biofilms with engineered enzymatic bacteriophage. Proceedings of the National Academy of Sciences of the United States of America 104, 11197-11202, doi:10.1073/pnas.0704624104 (2007).

6          Lemire, S., Yehl, K. M. & Lu, T. K. Phage-Based Applications in Synthetic Biology. Annual Review of Virology 5, 453-476, doi:10.1146/annurev-virology-092917-043544 (2018).