Definitions and guidelines for research on antibiotic persistence

Abstract

Increasing concerns about the rising rates of antibiotic therapy failure and advances in single-cell analyses have inspired a surge of research into antibiotic persistence. Bacterial persister cells represent a subpopulation of cells that can survive intensive antibiotic treatment without being resistant. Several approaches have emerged to define and measure persistence, and it is now time to agree on the basic definition of persistence and its relation to the other mechanisms by which bacteria survive exposure to bactericidal antibiotic treatments, such as antibiotic resistance, heteroresistance or tolerance. In this Consensus Statement, we provide definitions of persistence phenomena, distinguish between triggered and spontaneous persistence and provide a guide to measuring persistence. Antibiotic persistence is not only an interesting example of non-genetic single-cell heterogeneity, it may also have a role in the failure of antibiotic treatments. Therefore, it is our hope that the guidelines outlined in this article will pave the way for better characterization of antibiotic persistence and for understanding its relevance to clinical outcomes.

Fig. 1. Persistent infections versus antibiotic persistence.

Persistent infection is a general term to describe infections that are not efficiently cleared by the host, in contrast to the characteristic acute response that leads to clearance of many pathogens. Antibiotic persistence specifically describes the heterogeneous response of bacterial populations, in vitro or in the host, that results in a delayed clearance of the bacterial load by antibiotics.

Fig. 2. Antibiotic resistance, tolerance and persistence.

Resistance, tolerance and persistence are distinct responses to antibiotic treatment that lead to increased survival compared with susceptible cells. a | To inhibit the growth of resistant bacteria, a substantially higher minimum inhibitory concentration (MIC) of the antibiotic is needed than for susceptible bacteria. Notably, persistence and tolerance do not lead to an increase in the MIC compared with susceptible bacteria. b | By contrast, tolerance increases the minimum duration for killing (MDK; for example, for 99% of bacterial cells in the population (MDK₉₉)) compared with susceptible bacteria. c | Persistence leads to a similar MIC and a similar initial killing of the bacterial population compared with susceptible bacteria; however, the MDK for 99.99% of bacterial cells in the population (MDK_99.99) can be substantially higher owing to the survival of the persister cells. Note that pure exponential killing of the susceptible strain is rarely observed because most bacterial cultures have some level of persistence. The data shown are only illustrations and not actual measurements. Parts b and c are adapted with permission from ref., Springer Nature Limited (this material is excluded from the CC-BY-4.0 license).

Fig. 3. Triggered versus spontaneous persistence.

Triggered persistence requires a trigger for bacteria to become persisters (left). The persistence level will then depend on the intensity and duration of the trigger. For example, a common trigger for persistence is starvation. Even when the trigger is removed, persister bacteria may retain their phenotype for an extended duration. Spontaneous persistence occurs when the bacteria are in steady-state exponential growth (right). A fraction of the population switches stochastically to the persister phenotype at a rate that is constant during growth. Such steady-state conditions can be found in chemostats or serially diluted cultures, and care must be taken to ensure that the persisters do not originate from the inoculum or from the culture being too close to entry into the stationary phase.