Proximate and ultimate causes of the bactericidal action of antibiotics

Abstract

During the past 85 years of antibiotic use, we have learned a great deal about how these 'miracle' drugs work. We know the molecular structures and interactions of these drugs and their targets and the effects on the structure, physiology and replication of bacteria. Collectively, we know a great deal about these proximate mechanisms of action for virtually all antibiotics in current use. What we do not know is the ultimate mechanism of action; that is, how these drugs irreversibly terminate the 'individuality' of bacterial cells by removing barriers to the external world (cell envelopes) or by destroying their genetic identity (DNA). Antibiotics have many different 'mechanisms of action' that converge to irreversible lethal effects. In this Perspective, we consider what our knowledge of the proximate mechanisms of action of antibiotics and the pharmacodynamics of their interaction with bacteria tell us about the ultimate mechanisms by which these antibiotics kill bacteria.

Fig. 1. Killing rates of different antibiotics.

The findings of time–kill experiments measured as changes in the viable cell density of Staphylococcus aureus Newman exposed to 10 times the minimum inhibitory concentration of nine different antibiotics in Mueller–Hinton II medium at 37 °C with aeration are shown. Bacteriostatic antibiotics (left) stop growth and colony-forming units (CFUs) remain stable. By contrast, bactericidal antibiotics (right) kill bacteria and thus the CFUs drop. Of note, a resistant mutant emerged in the culture treated with rifampin, leading to resumed growth at the end of the experiment.

Fig. 2. Factors influencing the bactericidal activity of antibiotics in a susceptible bacterial cell.

Only free drug is active (1) and protein binding, for example to albumin or other plasma proteins, can reduce the level of available and thus active drug, as commonly observed for β-lactams. Uptake systems (such as porins) and barriers can prevent the drug from entering cells (2) or the drug is pumped out (3). Greater or weaker target-binding affinity also influences activity (4). The targeted function can also increase in the presence of the drug, thereby compensating for inhibition (5; for example, upregulation of RpoB by rifampin in mycobacteria). The target function involves the build-up of a cellular structure with slow turnover (for example, peptidoglycan), which increases the amount of time for the antibiotic to kill (6). The cells repair the damage produced by the drug (7), involving the SOS system. The bacteria have inducible antibiotic-deactivating mechanisms (8; for example, β-lactamases). The bacteria use alternative functions, bypassing those that are inhibited (9). Antibiotics differ in the extent to which they induce reactive oxygen species (ROS; deleterious) or SOS (potentially protective) responses (10). Low replication rates (involving the SOS system) reduce the killing activity (11). Activation of the RpoS-mediated stringent response produces a kind of ‘stationary phase’, reducing bactericidal potency (12). The two ultimate causes (bottom right) of a bactericidal effect are loss of spatial individuality by rupture of the limits with the environment (broken green line indicating disruption of the cell envelope) and loss of genetic individuality (broken blue line indicating disruption of the genome).

Fig. 3. Successive steps in the process of bacterial killing by antibiotics from six families.

These drugs (blue) directly interact with their targets (purple), which results in structural damage and/or quantitative or qualitative deficiencies of essential cell components. These changes, in turn, lead to envelope stress, DNA damage and/or the production of an excess of reactive oxygen species, which further contribute to the destructuring of cell membranes and nucleic acids. The net effect of these different processes (green) is the ultimate mechanism responsible for the loss of the cell’s individuality, its death (red).