Collective antibiotic tolerance: mechanisms, dynamics and intervention

Abstract

Bacteria have developed resistance against every antibiotic at a rate that is alarming considering the timescale at which new antibiotics are developed. Thus, there is a critical need to use antibiotics more effectively, extend the shelf life of existing antibiotics and minimize their side effects. This requires understanding the mechanisms underlying bacterial drug responses. Past studies have focused on survival in the presence of antibiotics by individual cells, as genetic mutants or persisters. Also important, however, is the fact that a population of bacterial cells can collectively survive antibiotic treatments lethal to individual cells. This tolerance can arise by diverse mechanisms, including resistance-conferring enzyme production, titration-mediated bistable growth inhibition, swarming and interpopulation interactions. These strategies can enable rapid population recovery after antibiotic treatment and provide a time window during which otherwise susceptible bacteria can acquire inheritable genetic resistance. Here, we emphasize the potential for targeting collective antibiotic tolerance behaviors as an antibacterial treatment strategy.

Figure 1. Bacterial survival modes

a. Bacterial survival modes can be at the individual or population level. Populations can use different mechanisms to survive antibiotic collectively. b. Collective antibiotic tolerance emerges when a population at a sufficiently high density can survive an antibiotic dose that would be lethal to a low density population. c. Initial cell density determines the outcome of a population treated by an antibiotic. For each antibiotic concentration, there is a critical initial density above which the population will recover; below this, the population will die.

Figure 2. [within Box 1]. Comparison of population level responses due to different forms of CAT or persistence

a. Time course simulations. After being treated with an antibiotic, the bacteria actively tolerating antibiotics through bistable ribosome inhibition, PCD, or QS recover much faster than persisters. b. Recovery times. For persisters and CAT bacteria alike, increasing the initial cell density decreases the time it takes for a population to recover from antibiotic treatment.

Figure 3. Underlying mechanisms of CAT

a. Antibiotic mediated altruistic death involves a subpopulation lysing to release effector proteins that benefit the remainder of the population. CAT can only emerge if the population is large enough to generate a collective effector protein concentration sufficient to inhibit the antibiotic before the antibiotic kills all of the population. b. Quorum sensing is often involved in regulating the expression of effector proteins or the maturation of biofilm formation, both of which can lead to CAT. c. Bistable inhibition of bacterial growth arises as a function of intracellular antibiotic concentration and bistable ribosome inhibition. Sufficiently dense populations can titrate the antibiotic such that the intracellular concentration does not inhibit ribosome synthesis. However, low density populations cannot titrate the antibiotic to a sublethal level, thus, ribosome synthesis will be inhibited. d. Multiple mechanisms can be used by a clonal population to survive antibiotic treatment. For example, swarming is a function of phenotype, QS, and PCD that confers antibiotic tolerance. e. Social interactions can facilitate the survival of multiple subpopulations (denoted by different colored cell outlines). For instance, one subpopulation can produce a signaling molecule necessary to upregulate a second subpopulation’s resistance mechanism and vice versa. There are also situations where one subpopulation donates a signaling molecule that protects a second population, without deriving any benefit in return.

Figure 4. Inhibition strategies

a. Timing control of antibiotic dosing is critical for optimizing efficacy of periodic antibiotic treatments. By timing the application of antibiotic doses with when the population is at a low density, the amount of public good and/or toxin released would be insufficient to allow the population to recover or harm the host. b. Molecular-level interventions of CAT could be used to trigger PCD and inhibit the production and/or the function of an effector protein.