The core goal of our project is to develop a strategy that would solve the problem of bacterial resistance to antibiotics once and for all. Our reasoning goes like this: while small molecules used as antibiotics cannot be easily evolved to overcome bacterial resistance when it occurs, bacteriophage can very much be evolved. Thus phage-based treatments could tackle the issue of resistance by a constant evolution of the treatment agent (phage) in line with bacteria. The most intriguing case would be if one could evolve an anti-phage against a bacterium from a patient in a sufficiently short time to cure the patient with the new phage! We therefore made a chemostat-based evolution system that generates a diversity of phage, selecting them on their ability to infect phage-resistant bacteria, and made a biobrick that could increase their rate of mutation.
A chematic of the workings of the phage evolution system.
The chemostat based evolution system consists of 3 chemostats. In the first, phage host bacteria are grown – these bacteria can be infected by the phage to be evolved. They are used to generate phage diversity and can host a mutagenic plasmid in order to make the diversity even greater. In the second chemostat resistant bacteria are grown on their own. The outputs from these two chemostats bring the host and resistant bacteria into a third chemostat where they mix together. The third chemostat also contains phage that are to be evolved. Neither of the input bacteria are subject to evolutionary pressure within their own chemostats. This and the high volume exchange rate inside this chemostat prevents phage-bacteria competitive ecosystems from evolving. If bacterial resistance to phage evolves in this chemostat, the resistant bacteria will get washed out and will not affect the evolution of phage.
Our real phage evolution system at work. The three chemostats are seen in the middle. The one to
the left grows resistant bacteria, the one to the right grows the hosts and in the third phage are evolved.
Phage versus Antibiotics
We used our chemostat system in different modes to first evolve a phage-resistant E.coli and then evolve
a phage to kill it within 13 hours of evolution. (See the results section for further details). This
showed that it is practical to perform fast phage evolution given a resistant bacterium. As compared
to the rate of invention of new antibiotics this is of course a lot faster. However it is also easier
for bacteria to evolve against phage than against antibiotics. A possible solution could be phage cocktails
or pluripotent phage. The phage mix that we evolved, for example, was able to kill both the original
bacterium and the one that was resistant to the original phage. In any case the system for allowing
new phage into the market would have to be more dynamic than that of antibiotic approval. This means
that the long times and huge amounts of money that are necessary to approve a new medicine would likely
need to be abolished in the case of phage. This will not happen unless legislative changes are introduced.
However, as the prevalence of bacterial resistance to antibiotics increases, we think that more pressure will be put on the authorities to pursue such changes.
There are already orders of magnitude more phage than bacteria in nature. The competition between bacteria and phage has lasted billions of years neither biological form being able to extinguish the other. It is therefore unlikely that we would manage to generate a phage to kill all bacteria in the lab given the time scales and the copy numbers that we are working with. It is likely that common pathogenic bacteria will evolve resistance against the phage used in treatment, but then we are back to the same issue that we successfully addressed in our project – we will always be able to evolve an antiphage.