With the number of antibiotic treatment failures rising at an alarming rate, we are in dire need of new anti-bacterial therapies. To discover new therapies, it is important to understand the mechanisms responsible for bacterial survival. We focussed on one survival strategy, antibiotic persistence, in which a subfraction of a bacterial population, referred to as persister cells, is in a temporarily antibiotic-tolerant state. In this tolerant state, persister cells do not grow and are therefore often seen as dormant. Under favourable conditions, these persister cells can recover from the antibiotic treatment and resume growth, thereby causing a relapse of the infection.
In this work, we aimed to identify mechanisms involved in the formation and recovery of persister cells. To this end, we explored a potential role in persistence for YbiB, a promising interaction partner of a previously identified persister gene. Furthermore, we developed a new high-throughput screening approach making use of CRISPRi to search for genes involved in persistence.
Our results show that YbiB is involved in persistence but the underlying cellular mechanisms remain to be discovered. Based on our CRISPRi results, we demonstrate that dormancy is not sufficient to survive antibiotic treatment. On the contrary, we find that persister survival depends on the downregulation of specific metabolic pathways. Furthermore, we demonstrate that persister cells need active mechanisms to withstand antibiotic treatment and that these mechanisms are dependent on the type of antibiotic used. For example, when persister cells acquire DNA damage by a DNA-damaging antibiotic, they need to repair this damage to survive during treatment but also to restart growth after treatment. Finally, we evaluated if persister cells are multidrug tolerant by treating them with a combination of two antibiotics. Our data confirm that some persister cells can survive this treatment and are thus multidrug tolerant, while others are killed. This indicates that persistence is a dynamic phenotype and that not all persister cells are equal.
Our research provides important insights in the mechanisms underlying persister formation and recovery. In the future, these results might serve as a basis for the development of anti-persister therapies.
